Anti-cd3 antibodies, activatable anti-cd3 antibodies, multispecific anti-cd3 antibodies, multispecific activatable anti-cd3 antibodies, and methods of using the same

ABSTRACT

The invention relates generally to antibodies, activatable antibodies, multispecific antibodies, and multispecific activatable antibodies that specifically bind to at least CD3, as well as to methods of making and using these antibodies, activatable antibodies, multispecific antibodies, and/or multispecific activatable antibodies in a variety of therapeutic, diagnostic and prophylactic indications.

RELATED APPLICATIONS

This application claims the benefit of U.S. Provisional Application No.62/029,325, filed Jul. 25, 2014, the contents of which are incorporatedherein by reference in their entirety.

FIELD OF THE DISCLOSURE

The invention relates generally to antibodies, activatable antibodies,multispecific antibodies, and multispecific activatable antibodies thatspecifically bind to at least CD3, as well as to methods of making andusing these antibodies, activatable antibodies, multispecificantibodies, and/or multispecific activatable antibodies in a variety oftherapeutic, diagnostic and prophylactic indications.

BACKGROUND OF THE DISCLOSURE

CD3 (Cluster of Differentiation 3) T-cell co-receptor is a multimericprotein composed of four distinct polypeptide chains, referred to as theε, γ, δ, and ζ chains. The CD3 complex serves as the signaling module ofthe T cell receptor that associates non-covalently with theantigen-binding a/b chains of T cell receptor (TCR).

Because direct engagement of CD3 results in T-cell activation, it is adesirable target for a variety of therapeutic and/or diagnosticindications. Accordingly, there exists a need for antibodies andtherapeutics that target the CD3/TCR pathway.

SUMMARY OF THE DISCLOSURE

The present disclosure provides antibodies and antigen-binding fragmentsthereof that specifically bind the epsilon chain of CD3, also known asCD3ε. The anti-CD3ε antibodies and antigen-binding fragments thereof ofthe disclosure activate T cells via engagement of CD3ε on the T cells.That is, such antibodies agonize, stimulate, activate, and/or augmentCD3-mediated T cell activation. These antibodies and antigen-bindingfragments thereof are referred to herein as “anti-CD3ε antibodies” or“anti-CD3 antibodies.” The anti-CD3ε antibodies and antigen-bindingfragments thereof of the disclosure include monoclonal antibodies, suchas, for example, mammalian monoclonal antibodies, primate monoclonalantibodies, fully human monoclonal antibodies, as well as humanizedmonoclonal antibodies and chimeric antibodies, as well asantigen-binding fragments thereof. In some embodiments, the antibodiesand antigen-binding fragments thereof are IgG isotype. In someembodiments, the antibodies and antigen-binding fragments thereof areIgG1 isotype. In some embodiments, the antibodies and antigen-bindingfragments thereof have one of any of the isotypes disclosed herein.

In some embodiments, the anti-CD3ε antibody or antigen-binding fragmentthereof includes a combination of a VH CDR1 sequence, a VH CDR2sequence, and a VH CDR3 sequence, wherein at least one of the VH CDR1sequence, the VH CDR2 sequence, and the VH CDR3 sequence is selectedfrom a VH CDR1 sequence that includes at least the amino acid sequenceTYAMN (SEQ ID NO: 53) a VH CD2 sequence that includes at least the aminoacid sequence RIRSKYNNYATYYADSVKD (SEQ ID NO: 54); and a VH CDR3sequence that includes at least the amino acid sequence HGNFGNSYVSWFAY(SEQ ID NO: 55).

In some embodiments, the anti-CD3ε antibody or antigen-binding fragmentthereof includes a combination of a VL CDR1 sequence, a VL CDR2sequence, and a VL CDR3 sequence, wherein at least one of the VL CDR1sequence, the VL CDR2 sequence, and the VL CDR3 sequence is selectedfrom a VL CDR1 sequence that includes at least the amino acid sequenceRSSTGAVTTSNYAN (SEQ ID NO: 56); a VL CDR2 sequence that includes atleast the amino acid sequence GTNKRAP (SEQ ID NO: 57); and a VL CDR3sequence that includes at least the amino acid sequence ALWYSNLWV (SEQID NO: 58).

In some embodiments, the anti-CD3ε antibody or antigen-binding fragmentthereof includes a combination of a VH CDR1 sequence, a VH CDR2sequence, and a VH CDR3 sequence, wherein at least one of the VH CDR1sequence, the VH CDR2 sequence, and the VH CDR3 sequence is selectedfrom a VH CDR1 sequence that includes at least the amino acid sequenceTYAMN (SEQ ID NO: 53); a VH CD2 sequence that includes at least theamino acid sequence RIRSKYNNYATYYADSVKD (SEQ ID NO: 54); a VH CDR3sequence that includes at least the amino acid sequence HGNFGNSYVSWFAY(SEQ ID NO: 55), and a combination of a VL CDR1 sequence, a VL CDR2sequence, and a VL CDR3 sequence, wherein at least one of the VL CDR1sequence, the VL CDR2 sequence, and the VL CDR3 sequence is selectedfrom a VL CDR1 sequence that includes at least the amino acid sequenceRSSTGAVTTSNYAN (SEQ ID NO: 56); a VL CDR2 sequence that includes atleast the amino acid sequence GTNKRAP (SEQ ID NO: 57); and a VL CDR3sequence that includes at least the amino acid sequence ALWYSNLWV (SEQID NO: 58).

In some embodiments, the anti-CD3ε antibody or antigen-binding fragmentthereof includes a VH CDR1 sequence that includes at least the aminoacid sequence TYAMN (SEQ ID NO: 53); a VH CD2 sequence that includes atleast the amino acid sequence RIRSKYNNYATYYADSVKD (SEQ ID NO: 54); a VHCDR3 sequence that includes at least the amino acid sequenceHGNFGNSYVSWFAY (SEQ ID NO: 55), a VL CDR1 sequence that includes atleast the amino acid sequence RSSTGAVTTSNYAN (SEQ ID NO: 56); a VL CDR2sequence that includes at least the amino acid sequence GTNKRAP (SEQ IDNO: 57); and a VL CDR3 sequence that includes at least the amino acidsequence ALWYSNLWV (SEQ ID NO: 58).

In some embodiments, the anti-CD3ε antibody or antigen-binding fragmentthereof includes a combination of a VH CDR1 sequence, a VH CDR2sequence, and a VH CDR3 sequence, wherein at least one of the VH CDR1sequence, the VH CDR2 sequence, and the VH CDR3 sequence is selectedfrom a VH CDR1 sequence that includes a sequence that is at least 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical to theamino acid sequence TYAMN (SEQ ID NO: 53); a VH CD2 sequence thatincludes a sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%,97%, 98%, 99% or more identical to the amino acid sequenceRIRSKYNNYATYYADSVKD (SEQ ID NO: 54); and a VH CDR3 sequence thatincludes a sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%,97%, 98%, 99% or more identical to the amino acid sequenceHGNFGNSYVSWFAY (SEQ ID NO: 55).

In some embodiments, the anti-CD3ε antibody or antigen-binding fragmentthereof includes a combination of a VL CDR1 sequence, a VL CDR2sequence, and a VL CDR3 sequence, wherein at least one of the VL CDR1sequence, the VL CDR2 sequence, and the VL CDR3 sequence is selectedfrom a VL CDR1 sequence that includes a sequence that is at least 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical to theamino acid sequence RSSTGAVTTSNYAN (SEQ ID NO: 56); a VL CDR2 sequencethat includes a sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%,96%, 97%, 98%, 99% or more identical to the amino acid sequence GTNKRAP(SEQ ID NO: 57); and a VL CDR3 sequence that includes a sequence that isat least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or moreidentical to the amino acid sequence ALWYSNLWV (SEQ ID NO: 58).

In some embodiments, the anti-CD3ε antibody or antigen-binding fragmentthereof includes a combination of a VH CDR1 sequence, a VH CDR2sequence, and a VH CDR3 sequence, wherein at least one of the VH CDR1sequence, the VH CDR2 sequence, and the VH CDR3 sequence is selectedfrom a VH CDR1 sequence that includes a sequence that is at least 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical to theamino acid sequence TYAMN (SEQ ID NO: 53); a VH CD2 sequence thatincludes a sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%,97%, 98%, 99% or more identical to the amino acid sequenceRIRSKYNNYATYYADSVKD (SEQ ID NO: 54); a VH CDR3 sequence that includes asequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%,99% or more identical to the amino acid sequence HGNFGNSYVSWFAY (SEQ IDNO: 55), and a combination of a VL CDR1 sequence, a VL CDR2 sequence,and a VL CDR3 sequence, wherein at least one of the VL CDR1 sequence,the VL CDR2 sequence, and the VL CDR3 sequence is selected from a VLCDR1 sequence that includes a sequence that is at least 90%, 91%, 92%,93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical to the amino acidsequence RSSTGAVTTSNYAN (SEQ ID NO: 56); a VL CDR2 sequence thatincludes a sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%,97%, 98%, 99% or more identical to the amino acid sequence GTNKRAP (SEQID NO: 57); and a VL CDR3 sequence that includes a sequence that is atleast 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identicalto the amino acid sequence ALWYSNLWV (SEQ ID NO: 58).

In some embodiments, the anti-CD3ε antibody or antigen-binding fragmentthereof includes a VH CDR1 sequence that is at least 90%, 91%, 92%, 93%,94%, 95%, 96%, 97%, 98%, 99% or more identical to the amino acidsequence TYAMN (SEQ ID NO: 53); a VH CD2 sequence that is at least 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical to theamino acid sequence RIRSKYNNYATYYADSVKD (SEQ ID NO: 54); a VH CDR3sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%,99% or more identical to the amino acid sequence HGNFGNSYVSWFAY (SEQ IDNO: 55), a VL CDR1 sequence that is at least 90%, 91%, 92%, 93%, 94%,95%, 96%, 97%, 98%, 99% or more identical to the amino acid sequenceRSSTGAVTTSNYAN (SEQ ID NO: 56); a VL CDR2 sequence that is at least 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical to theamino acid sequence GTNKRAP (SEQ ID NO: 57); and a VL CDR3 sequence thatis at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or moreidentical to the amino acid sequence ALWYSNLWV (SEQ ID NO: 58).

In some embodiments, the anti-CD3ε antibody or antigen-binding fragmentthereof includes a variable heavy chain (Hv) comprising the amino acidsequence of SEQ ID NO: 4. In some embodiments, the anti-CD3ε antibody orantigen-binding fragment thereof includes a variable light chain (Lv)comprising the amino acid sequence of SEQ ID NO: 2. In some embodiments,the anti-CD3ε antibody or antigen-binding fragment thereof includes avariable heavy chain (Hv) comprising the amino acid sequence of SEQ IDNO: 4 and a variable light chain (Lv) comprising the amino acid sequenceof SEQ ID NO: 2.

In some embodiments, the anti-CD3ε antibody or antigen-binding fragmentthereof includes a variable heavy chain (Hv) comprising an amino acidsequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%,99% or more identical to the amino acid sequence of SEQ ID NO: 4. Insome embodiments, the anti-CD3ε antibody or antigen-binding fragmentthereof includes a variable light chain (Lv) comprising an amino acidsequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%,99% or more identical to the amino acid sequence of SEQ ID NO: 2. Insome embodiments, the anti-CD3ε antibody or antigen-binding fragmentthereof includes a variable heavy chain (Hv) comprising an amino acidsequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%,99% or more identical to the amino acid sequence of SEQ ID NO: 4 and avariable light chain (Lv) comprising the amino acid sequence of SEQ IDNO: 2.

In some embodiments, the anti-CD3ε antibody is an scFv antibody fragmentthat binds CD3ε. In some embodiments, the anti-CD3ε scFv antibodyfragment includes the amino acid sequence of SEQ ID NO: 6. In someembodiments, the anti-CD3ε scFv antibody fragment includes the aminoacid sequence of SEQ ID NO: 30. In some embodiments, the anti-CD3ε scFvantibody fragment includes an amino acid sequence that is at least 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical to theamino acid sequence of SEQ ID NO: 6. In some embodiments, the anti-CD3εscFv antibody fragment includes an amino acid sequence that is at least90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical tothe amino acid sequence of SEQ ID NO: 30.

In some embodiments, the anti-CD3ε antibody also includes an agentconjugated to the antibody. In some embodiments, the agent is atherapeutic agent. In some embodiments, the agent is a detectablemoiety. In some embodiments, the detectable moiety is a diagnosticagent. In some embodiments, the agent is conjugated to the anti-CD3εantibody via a linker. In some embodiments, the linker is a cleavablelinker. In some embodiments, the linker is a non-cleavable linker.

In some embodiments, the anti-CD3ε antibody naturally contains one ormore disulfide bonds. In some embodiments, the anti-CD3ε antibody can beengineered to include one or more disulfide bonds.

The disclosure also provides an isolated nucleic acid molecule encodingat least a portion of an anti-CD3ε antibody described herein and/or oneor more nucleic acid molecules encoding an anti-CD3ε antibody describedherein, such as for example, at least a first nucleic acid encoding atleast a portion of the heavy chain of the antibody and a second nucleicacid encoding at least a portion of the light chain of the antibody, aswell as vectors that include these isolated nucleic acid sequences. Thedisclosure provides methods of producing an anti-CD3ε antibody byculturing a cell under conditions that lead to expression of theanti-CD3ε antibody, wherein the cell comprises such a nucleic acidmolecule(s). In some embodiments, the cell comprises such a vector.

The disclosure also provides activatable antibodies and activatableantibody compositions that include an antibody or antigen-bindingfragment thereof (AB) that specifically binds CD3ε coupled or otherwiseattached to a masking moiety (MM), such that coupling of the MM reducesthe ability of the AB to bind CD3ε. These activatable antibodies arecollectively referred to herein as activatable anti-CD3ε antibodies,also referred to herein as anti-CD3ε activatable antibodies or CD3εactivatable antibodies. In some embodiments, the MM is coupled via asequence that includes a substrate for a protease. For example, theprotease is produced by a tumor that is in proximity to cells thatexpress CD3ε. In some embodiments, the protease is produced by a tumorthat is co-localized with cells that express CD3ε. The activatableanti-CD3ε antibodies provided herein are stable in circulation,activated at intended sites of therapy and/or diagnosis but not innormal, i.e., healthy tissue, and, when activated, exhibit binding toCD3ε that is at least comparable to the corresponding, unmodifiedantibody.

In some embodiments, the activatable anti-CD3ε antibody orantigen-binding fragment thereof includes a combination of a VH CDR1sequence, a VH CDR2 sequence, and a VH CDR3 sequence, wherein at leastone of the VH CDR1 sequence, the VH CDR2 sequence, and the VH CDR3sequence is selected from a VH CDR1 sequence that includes at least theamino acid sequence TYAMN (SEQ ID NO: 53); a VH CD2 sequence thatincludes at least the amino acid sequence RIRSKYNNYATYYADSVKD (SEQ IDNO: 54); and a VH CDR3 sequence that includes at least the amino acidsequence HGNFGNSYVSWFAY (SEQ ID NO: 55).

In some embodiments, the activatable anti-CD3ε antibody orantigen-binding fragment thereof includes a combination of a VL CDR1sequence, a VL CDR2 sequence, and a VL CDR3 sequence, wherein at leastone of the VL CDR1 sequence, the VL CDR2 sequence, and the VL CDR3sequence is selected from a VL CDR1 sequence that includes at least theamino acid sequence RSSTGAVTTSNYAN (SEQ ID NO: 56); a VL CDR2 sequencethat includes at least the amino acid sequence GTNKRAP (SEQ ID NO: 57);and a VL CDR3 sequence that includes at least the amino acid sequenceALWYSNLWV (SEQ ID NO: 58).

In some embodiments, the activatable anti-CD3ε antibody orantigen-binding fragment thereof includes a combination of a VH CDR1sequence, a VH CDR2 sequence, and a VH CDR3 sequence, wherein at leastone of the VH CDR1 sequence, the VH CDR2 sequence, and the VH CDR3sequence is selected from a VH CDR1 sequence that includes at least theamino acid sequence TYAMN (SEQ ID NO: 53); a VH CD2 sequence thatincludes at least the amino acid sequence RIRSKYNNYATYYADSVKD (SEQ IDNO: 54); a VH CDR3 sequence that includes at least the amino acidsequence HGNFGNSYVSWFAY (SEQ ID NO: 55), and a combination of a VL CDR1sequence, a VL CDR2 sequence, and a VL CDR3 sequence, wherein at leastone of the VL CDR1 sequence, the VL CDR2 sequence, and the VL CDR3sequence is selected from a VL CDR1 sequence that includes at least theamino acid sequence RSSTGAVTTSNYAN (SEQ ID NO: 56); a VL CDR2 sequencethat includes at least the amino acid sequence GTNKRAP (SEQ ID NO: 57);and a VL CDR3 sequence that includes at least the amino acid sequenceALWYSNLWV (SEQ ID NO: 58).

In some embodiments, the activatable anti-CD3ε antibody orantigen-binding fragment thereof includes a VH CDR1 sequence thatincludes at least the amino acid sequence TYAMN (SEQ ID NO: 53); a VHCD2 sequence that includes at least the amino acid sequenceRIRSKYNNYATYYADSVKD (SEQ ID NO: 54); a VH CDR3 sequence that includes atleast the amino acid sequence HGNFGNSYVSWFAY (SEQ ID NO: 55), a VL CDR1sequence that includes at least the amino acid sequence RSSTGAVTTSNYAN(SEQ ID NO: 56); a VL CDR2 sequence that includes at least the aminoacid sequence GTNKRAP (SEQ ID NO: 57); and a VL CDR3 sequence thatincludes at least the amino acid sequence ALWYSNLWV (SEQ ID NO: 58).

In some embodiments, the activatable anti-CD3ε antibody orantigen-binding fragment thereof includes a combination of a VH CDR1sequence, a VH CDR2 sequence, and a VH CDR3 sequence, wherein at leastone of the VH CDR1 sequence, the VH CDR2 sequence, and the VH CDR3sequence is selected from a VH CDR1 sequence that includes a sequencethat is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% ormore identical to the amino acid sequence TYAMN (SEQ ID NO: 53); a VHCD2 sequence that includes a sequence that is at least 90%, 91%, 92%,93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical to the amino acidsequence RIRSKYNNYATYYADSVKD (SEQ ID NO: 54); and a VH CDR3 sequencethat includes a sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%,96%, 97%, 98%, 99% or more identical to the amino acid sequenceHGNFGNSYVSWFAY (SEQ ID NO: 55).

In some embodiments, the activatable anti-CD3ε antibody orantigen-binding fragment thereof includes a combination of a VL CDR1sequence, a VL CDR2 sequence, and a VL CDR3 sequence, wherein at leastone of the VL CDR1 sequence, the VL CDR2 sequence, and the VL CDR3sequence is selected from a VL CDR1 sequence that includes a sequencethat is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% ormore identical to the amino acid sequence RSSTGAVTTSNYAN (SEQ ID NO:56); a VL CDR2 sequence that includes a sequence that is at least 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical to theamino acid sequence GTNKRAP (SEQ ID NO: 57); and a VL CDR3 sequence thatincludes a sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%,97%, 98%, 99% or more identical to the amino acid sequence ALWYSNLWV(SEQ ID NO: 58).

In some embodiments, the activatable anti-CD3ε antibody orantigen-binding fragment thereof includes a combination of a VH CDR1sequence, a VH CDR2 sequence, and a VH CDR3 sequence, wherein at leastone of the VH CDR1 sequence, the VH CDR2 sequence, and the VH CDR3sequence is selected from a VH CDR1 sequence that includes a sequencethat is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% ormore identical to the amino acid sequence TYAMN (SEQ ID NO: 53); a VHCD2 sequence that includes a sequence that is at least 90%, 91%, 92%,93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical to the amino acidsequence RIRSKYNNYATYYADSVKD (SEQ ID NO: 54); a VH CDR3 sequence thatincludes a sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%,97%, 98%, 99% or more identical to the amino acid sequenceHGNFGNSYVSWFAY (SEQ ID NO: 55), and a combination of a VL CDR1 sequence,a VL CDR2 sequence, and a VL CDR3 sequence, wherein at least one of theVL CDR1 sequence, the VL CDR2 sequence, and the VL CDR3 sequence isselected from a VL CDR1 sequence that includes a sequence that is atleast 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identicalto the amino acid sequence RSSTGAVTTSNYAN (SEQ ID NO: 56); a VL CDR2sequence that includes a sequence that is at least 90%, 91%, 92%, 93%,94%, 95%, 96%, 97%, 98%, 99% or more identical to the amino acidsequence GTNKRAP (SEQ ID NO: 57); and a VL CDR3 sequence that includes asequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%,99% or more identical to the amino acid sequence ALWYSNLWV (SEQ ID NO:58).

In some embodiments, the activatable anti-CD3ε antibody orantigen-binding fragment thereof includes a VH CDR1 sequence that is atleast 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identicalto the amino acid sequence TYAMN (SEQ ID NO: 53); a VH CD2 sequence thatis at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or moreidentical to the amino acid sequence RIRSKYNNYATYYADSVKD (SEQ ID NO:54); a VH CDR3 sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%,96%, 97%, 98%, 99% or more identical to the amino acid sequenceHGNFGNSYVSWFAY (SEQ ID NO: 55), a VL CDR1 sequence that is at least 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical to theamino acid sequence RSSTGAVTTSNYAN (SEQ ID NO: 56); a VL CDR2 sequencethat is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% ormore identical to the amino acid sequence GTNKRAP (SEQ ID NO: 57); and aVL CDR3 sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%,97%, 98%, 99% or more identical to the amino acid sequence ALWYSNLWV(SEQ ID NO: 58).

In some embodiments, the activatable antibody comprises a combination ofa variable heavy chain complementarity determining region 1 (VH CDR1,also referred to herein as CDRH1) sequence, a variable heavy chaincomplementarity determining region 2 (VH CDR2, also referred to hereinas CDRH2) sequence, a variable heavy chain complementarity determiningregion 3 (VH CDR3, also referred to herein as CDRH3) sequence, avariable light chain complementarity determining region 1 (VL CDR1, alsoreferred to herein as CDRL1) sequence, a variable light chaincomplementarity determining region 2 (VL CDR2, also referred to hereinas CDRL2) sequence, and a variable light chain complementaritydetermining region 3 (VL CDR3, also referred to herein as CDRL3)sequence, wherein at least one CD3ε sequence is selected from the groupconsisting of a VH CDR1 sequence shown in Table 18; a VH CDR2 sequenceshown in Table 18; a VH CDR3 sequence shown in Table 18; a VL CDR1sequence shown in Table 18; a VL CDR2 sequence shown in Table 18; and aVL CDR3 sequence shown in Table 18.

In some embodiments, the activatable antibody comprises a combination ofa VH CDR1 sequence, a VH CDR2 sequence, a VH CDR3 sequence, a VL CDR1sequence, a VL CDR2 sequence, and a VL CDR3 sequence, wherein at leastone CD3ε sequence is selected from the group consisting of a VH CDR1sequence that includes a sequence that is at least 90%, 91%, 92%, 93%,94%, 95%, 96%, 97%, 98%, 99% or more identical to a VH CDR1 sequenceshown in Table 18; a VH CD2 sequence that includes a sequence that is atleast 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identicalto a VH CDR2 sequence shown in Table 18; a VH CDR3 sequence thatincludes a sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%,97%, 98%, 99% or more identical to a VH CDR3 sequence shown in Table 18;a VL CDR1 sequence that includes a sequence that is at least 90%, 91%,92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical to a VL CDR1sequence shown in Table 18; a VL CDR2 sequence that includes a sequencethat is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% ormore identical to a VL CDR2 sequence shown in Table 18; and a VL CDR3sequence that includes a sequence that is at least 90%, 91%, 92%, 93%,94%, 95%, 96%, 97%, 98%, 99% or more identical to a VL CDR3 sequenceshown in Table 18.

In some embodiments, the activatable antibody comprises a combination ofa VH CDR1 sequence, a VH CDR2 sequence, a VH CDR3 sequence, a VL CDR1sequence, a VL CDR2 sequence, and a VL CDR3 sequence, wherein thecombination is a combination shown in Table 18.

In some embodiments, the activatable antibody comprises a combination ofa VH CDR1 sequence, a VH CDR2 sequence, a VH CDR3 sequence, a VL CDR1sequence, a VL CDR2 sequence, and a VL CDR3 sequence, wherein each CD3εsequence in the combination comprises a sequence that is at least 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical to thecorresponding CD3ε sequence in a combination shown in Table 18.

In some embodiments, the activatable anti-CD3ε antibody orantigen-binding fragment thereof includes a variable heavy chain (Hv)comprising the amino acid sequence of SEQ ID NO: 4. In some embodiments,the anti-CD3ε antibody or antigen-binding fragment thereof includes avariable light chain (Lv) comprising the amino acid sequence of SEQ IDNO: 2. In some embodiments, the anti-CD3ε antibody or antigen-bindingfragment thereof includes a variable heavy chain (Hv) comprising theamino acid sequence of SEQ ID NO: 4 and a variable light chain (Lv)comprising the amino acid sequence of SEQ ID NO: 2.

In some embodiments, the activatable anti-CD3ε antibody orantigen-binding fragment thereof includes a variable heavy chain (Hv)comprising an amino acid sequence that is at least 90%, 91%, 92%, 93%,94%, 95%, 96%, 97%, 98%, 99% or more identical to the amino acidsequence of SEQ ID NO: 4. In some embodiments, the anti-CD3ε antibody orantigen-binding fragment thereof includes a variable light chain (Lv)comprising an amino acid sequence that is at least 90%, 91%, 92%, 93%,94%, 95%, 96%, 97%, 98%, 99% or more identical to the amino acidsequence of SEQ ID NO: 2. In some embodiments, the anti-CD3ε antibody orantigen-binding fragment thereof includes a variable heavy chain (Hv)comprising an amino acid sequence that is at least 90%, 91%, 92%, 93%,94%, 95%, 96%, 97%, 98%, 99% or more identical to the amino acidsequence of SEQ ID NO: 4 and a variable light chain (Lv) comprising theamino acid sequence of SEQ ID NO: 2.

In some embodiments, the activatable anti-CD3ε antibody includes an scFvantibody fragment that binds CD3ε. In some embodiments, the anti-CD3εscFv antibody fragment includes the amino acid sequence of SEQ ID NO: 6.In some embodiments, the anti-CD3ε scFv antibody fragment includes theamino acid sequence of SEQ ID NO: 30. In some embodiments, the anti-CD3εscFv antibody fragment includes an amino acid sequence that is at least90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical tothe amino acid sequence of SEQ ID NO: 6. In some embodiments, theanti-CD3ε scFv antibody fragment includes an amino acid sequence that isat least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or moreidentical to the amino acid sequence of SEQ ID NO: 30.

In some embodiments, the activatable antibody comprises a heavy chainamino acid sequence selected from the group consisting of the heavychain sequences shown in Table 17. In some embodiments, the activatableantibody comprises a light chain amino acid sequence selected from thegroup consisting of the light chain sequences shown in Table 17. In someembodiments, the activatable antibody comprises a heavy chain amino acidsequence selected from the group consisting of the heavy chain sequencesshown in Table 17 and a light chain amino acid sequence selected fromthe group consisting of the light chain sequences shown in Table 17.

In some embodiments, the activatable antibody comprises a heavy chainamino acid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%,97%, 98% or 99% identical to an amino acid sequence selected from thegroup consisting of the heavy chain sequences shown in Table 17. In someembodiments, the activatable antibody comprises a light chain amino acidsequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or99% identical to an amino acid sequence selected from the groupconsisting of the light chain sequences shown in Table 17. In someembodiments, the activatable antibody comprises a heavy chain amino acidsequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or99% identical to an amino acid sequence selected from the groupconsisting of the heavy chain sequences shown in Table 17 and a lightchain amino acid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%,96%, 97%, 98% or 99% identical to an amino acid sequence selected fromthe group consisting of the light chain sequences shown in Table 17.

In some embodiments, the MM has a dissociation constant, i.e.,dissociation constant at an equilibrium state, K_(d) for binding to theAB that is greater than the K_(d) for binding of the AB to CD3ε.

In some embodiments, the MM has a K_(d) for binding to the AB that is nomore than the K_(d) for binding of the AB to CD3ε.

In some embodiments, the MM has a K_(d) for binding to the AB that is noless than the K_(d) for binding of the AB to CD3ε.

In some embodiments, the MM has a K_(d) for binding to the AB that isapproximately equal to the K_(d) for binding of the AB to CD3ε.

In some embodiments, the MM has a K_(d) for binding to the AB that isless than the IQ for binding of the AB to CD3ε.

In some embodiments, the MM has a K_(d) for binding to the AB that is nomore than 2, 3, 4, 5, 10, 25, 50, 100, 250, 500, or 1,000 fold greaterthan the K_(d) for binding of the AB to CD3ε. In some embodiments, theMM has a K_(d) for binding to the AB that is between 1-5, 2-5, 2-10,5-10, 5-20, 5-50, 5-100, 10-100, 10-1,000, 20-100, 20-1000, or 100-1,000fold greater than the K_(d) for binding of the AB to CD3ε.

In some embodiments, the MM has an affinity for binding to the AB thatis less than the affinity of binding of the AB to CD3ε.

In some embodiments, the MM has an affinity for binding to the AB thatis no more than the affinity of binding of the AB to CD3ε.

In some embodiments, the MM has an affinity for binding to the AB thatis approximately equal of the affinity of binding of the AB to CD3ε.

In some embodiments, the MM has an affinity for binding to the AB thatis no less than the affinity of binding of the AB to CD3ε.

In some embodiments, the MM has an affinity for binding to the AB thatis greater than the affinity of binding of the AB to CD3ε.

In some embodiments, the MM has an affinity for binding to the AB thatis 2, 3, 4, 5, 10, 25, 50, 100, 250, 500, or 1,000 less than theaffinity of binding of the AB to CD3ε. In some embodiments, the MM hasan affinity for binding to the AB that is between 1-5, 2-5, 2-10, 5-10,5-20, 5-50, 5-100, 10-100, 10-1,000, 20-100, 20-1000, or 100-1,000 foldless than the affinity of binding of the AB to CD3ε. In someembodiments, the MM has an affinity for binding to the AB that is 2 to20 fold less than the affinity of binding of the AB to CD3ε. In someembodiments, a MM not covalently linked to the AB and at equimolarconcentration to the AB does not inhibit the binding of the AB to CD3ε.

In some embodiments, the MM does not interfere or compete with the ABfor binding to CD3ε when the activatable antibody is in a cleaved state.

In some embodiments, the MM is a polypeptide of no more than 40 aminoacids in length.

In some embodiments, the MM polypeptide sequence is different from thatof CD3ε. In some embodiments, the MM polypeptide sequence is no morethan 50% identical to any natural binding partner of the AB. In someembodiments, the MM polypeptide sequence is no more than 25% identicalto any natural binding partner of the AB. In some embodiments, the MMpolypeptide sequence is no more than 10% identical to any naturalbinding partner of the AB.

In some embodiments, the MM comprises a sequence selected from the groupconsisting of the sequences shown in Table 7 or Table 8.

In some embodiments, the protease is produced by a tumor that is inproximity to cells that express CD3ε and/or is produced by a tumor thatis co-localized with cells that express CD3ε in a tissue, and whereinthe protease cleaves the CM in the activatable antibody when theactivatable antibody is exposed to the protease. In some embodiments,the CM is a polypeptide of up to 15 amino acids in length. In someembodiments, the CM is a substrate for a protease selected from theproteases listed in Table 3. In some embodiments, the CM is a substratefor a protease selected from the group consisting of uPA, legumain,matriptase (also referred to herein as MT-SP1 or MTSP1), ADAM17, BMP-1,TMPRSS3, TMPRSS4, MMP-9, MMP-12, MMP-13, MMP-14, and any of those shownin Table 3. In some embodiments, the CM is a substrate for an proteaseselected from the group consisting of uPA, legumain, and matriptase. Insome embodiments, the CM is a substrate for a matrix metalloprotease(MMP).

In some embodiments, the antigen binding fragment thereof is selectedfrom the group consisting of a Fab fragment, a F(ab′)₂ fragment, a scFv,a scab, a dAb, a single domain heavy chain antibody, and a single domainlight chain antibody.

In some embodiments, the activatable antibody in the uncleaved state hasthe structural arrangement from N-terminus to C-terminus as follows:MM-CM-AB or AB-CM-MM. In some embodiments, the activatable antibodycomprises a linking peptide between the MM and the CM. In someembodiments, the activatable antibody comprises a linking peptidebetween the CM and the AB. In some embodiments, the activatable antibodycomprises a first linking peptide (LP1) and a second linking peptide(LP2), wherein the activatable antibody in the uncleaved state has thestructural arrangement from N-terminus to C-terminus as follows:MM-LP1-CM-LP2-AB or AB-LP2-CM-LP1-MM. In some embodiments, the twolinking peptides need not be identical to each other. In someembodiments, each of LP1 and LP2 is a peptide of about 1 to 20 aminoacids in length.

In some embodiments, the activatable anti-CD3ε antibody also includes anagent conjugated to the activatable antibody. In some embodiments, theagent is a therapeutic agent. In some embodiments, the agent is adetectable moiety. In some embodiments, the detectable moiety is adiagnostic agent. In some embodiments, the agent is conjugated to theactivatable anti-CD3ε antibody via a linker. In some embodiments, thelinker is a cleavable linker. In some embodiments, the linker is anon-cleavable linker.

The disclosure also provides multispecific antibodies that bind theepsilon chain of CD3 (CD3ε) and a second target, wherein the antibodycomprises a first antibody or antigen-binding fragment thereof (AB1)that binds the epsilon chain of CD3 (CD3ε) and a second antibody orantigen-binding fragment thereof (AB2) that binds a second target, andwherein AB1 comprises a VH CDR1 sequence that includes at least theamino acid sequence TYAMN (SEQ ID NO: 53); a VH CD2 sequence thatincludes at least the amino acid sequence RIRSKYNNYATYYADSVKD (SEQ IDNO: 54); a VH CDR3 sequence that includes at least the amino acidsequence HGNFGNSYVSWFAY (SEQ ID NO: 55), and a combination of a VL CDR1sequence, a VL CDR2 sequence, and a VL CDR3 sequence, wherein at leastone of the VL CDR1 sequence, the VL CDR2 sequence, and the VL CDR3sequence is selected from a VL CDR1 sequence that includes at least theamino acid sequence RSSTGAVTTSNYAN (SEQ ID NO: 56); a VL CDR2 sequencethat includes at least the amino acid sequence GTNKRAP (SEQ ID NO: 57);and a VL CDR3 sequence that includes at least the amino acid sequenceALWYSNLWV (SEQ ID NO: 58).

In some embodiments, the activatable antibodies described herein in anactivated state bind CD3ε and include (i) an antibody or an antigenbinding fragment thereof (AB) that specifically binds to CD3ε; (ii) amasking moiety (MM) that inhibits the binding of the AB to CD3ε when theactivatable antibody is in an uncleaved state; and (c) a cleavablemoiety (CM) coupled to the AB, wherein the CM is a polypeptide thatfunctions as a substrate for a protease. In some embodiments, the MM iscoupled to the AB via the CM.

In some embodiments, the activatable antibody in the uncleaved state hasthe structural arrangement from N-terminus to C-terminus as follows:MM-CM-AB or AB-CM-MM.

In some embodiments, the activatable antibody comprises a linkingpeptide between the MM and the CM.

In some embodiments, the activatable antibody comprises a linkingpeptide between the CM and the AB.

In some embodiments, the activatable antibody comprises a first linkingpeptide (LP1) and a second linking peptide (LP2), and the activatableantibody in the uncleaved state has the structural arrangement fromN-terminus to C-terminus as follows: MM-LP1-CM-LP2-AB orAB-LP2-CM-LP1-MM. In some embodiments, the two linking peptides need notbe identical to each other.

In some embodiments, at least one of LP1 or LP2 includes an amino acidsequence selected from the group consisting of (GS)_(n), (GGS)_(n),(GSGGS)_(n) (SEQ ID NO: 59) and (GGGS)_(n) (SEQ ID NO: 60), where n isan integer of at least one. In some embodiments, at least one of LP1 orLP2 includes an amino acid sequence selected from the group consistingof GGSG (SEQ ID NO: 61), GGSGG (SEQ ID NO: 62), GSGSG (SEQ ID NO: 63),GSGGG (SEQ ID NO: 64), GGGSG (SEQ ID NO: 65), and GSSSG (SEQ ID NO: 66).

In some embodiments, the activatable antibody includes an antibody orantigen-binding fragment thereof that specifically binds CD3ε. In someembodiments, the antibody or antigen-binding fragment thereof that bindsCD3ε is a monoclonal antibody, domain antibody, single chain, Fabfragment, a F(ab′)₂ fragment, a scFv, a scAb, a dAb, a single domainheavy chain antibody, or a single domain light chain antibody. In someembodiments, such an antibody or antigen-binding fragment thereof thatbinds CD3ε is a rodent (e.g., mouse or rat), chimeric, humanized orfully human monoclonal antibody.

In some embodiments, the AB has a dissociation constant of about 100 nMor less for binding to CD3ε.

In some embodiments, the MM has a dissociation constant, i.e.,dissociation constant at an equilibrium state, K_(d) for binding to theAB that is greater than the K_(d) for binding of the AB to CD3ε.

In some embodiments, the MM has a K_(d) for binding to the AB that is nomore than the K_(d) for binding of the AB to CD3ε.

In some embodiments, the MM has a K_(d) for binding to the AB that is noless than the K_(d) for binding of the AB to CD3ε.

In some embodiments, the MM has a K_(d) for binding to the AB that isapproximately equal to the K_(d) for binding of the AB to CD3ε.

In some embodiments, the MM has a K_(d) for binding to the AB that isless than the IQ for binding of the AB to CD3ε.

In some embodiments, the MM has a K_(d) for binding to the AB that is nomore than 2, 3, 4, 5, 10, 25, 50, 100, 250, 500, or 1,000 fold greaterthan the K_(d) for binding of the AB to CD3ε. In some embodiments, theMM has a K_(d) for binding to the AB that is between 1-5, 2-5, 2-10,5-10, 5-20, 5-50, 5-100, 10-100, 10-1,000, 20-100, 20-1000, or 100-1,000fold greater than the K_(d) for binding of the AB to CD3ε.

In some embodiments, the MM has an affinity for binding to the AB thatis less than the affinity of binding of the AB to CD3ε.

In some embodiments, the MM has an affinity for binding to the AB thatis no more than the affinity of binding of the AB to CD3ε.

In some embodiments, the MM has an affinity for binding to the AB thatis approximately equal of the affinity of binding of the AB to CD3ε.

In some embodiments, the MM has an affinity for binding to the AB thatis no less than the affinity of binding of the AB to CD3ε.

In some embodiments, the MM has an affinity for binding to the AB thatis greater than the affinity of binding of the AB to CD3ε.

In some embodiments, the MM has an affinity for binding to the AB thatis 2, 3, 4, 5, 10, 25, 50, 100, 250, 500, or 1,000 less than theaffinity of binding of the AB to CD3ε. In some embodiments, the MM hasan affinity for binding to the AB that is between 1-5, 2-5, 2-10, 5-10,5-20, 5-50, 5-100, 10-100, 10-1,000, 20-100, 20-1000, or 100-1,000 foldless than the affinity of binding of the AB to CD3ε. In someembodiments, the MM has an affinity for binding to the AB that is 2 to20 fold less than the affinity of binding of the AB to CD3ε. In someembodiments, a MM not covalently linked to the AB and at equimolarconcentration to the AB does not inhibit the binding of the AB to CD3ε.

In some embodiments, the MM does not interfere or compete with the ABfor binding to CD3ε when the activatable antibody is in a cleaved state.

In some embodiments, the MM is a polypeptide of about 2 to 40 aminoacids in length. In some embodiments, the MM is a polypeptide of no morethan 40 amino acids in length.

In some embodiments, the MM polypeptide sequence is different from thatof CD3ε. In some embodiments, the MM polypeptide sequence is no morethan 50% identical to any natural binding partner of the AB. In someembodiments, the MM polypeptide sequence is different from that of CD3εand wherein the MM polypeptide sequence is no more than 25% identical toany natural binding partner of the AB. In some embodiments, the MMpolypeptide sequence is different from that of CD3ε and wherein the MMpolypeptide sequence is no more than 10% identical to any naturalbinding partner of the AB.

In some embodiments, the coupling of the MM reduces the ability of theAB to bind CD3ε such that the dissociation constant (K_(d)) of the ABwhen coupled to the MM towards CD3ε is at least 20 times greater thanthe K_(d) of the AB when not coupled to the MM towards CD3ε.

In some embodiments, the coupling of the MM reduces the ability of theAB to bind CD3ε such that the dissociation constant (K_(d)) of the ABwhen coupled to the MM towards CD3ε is at least 40 times greater thanthe K_(d) of the AB when not coupled to the MM towards CD3ε.

In some embodiments, the coupling of the MM reduces the ability of theAB to bind CD3ε such that the dissociation constant (K_(d)) of the ABwhen coupled to the MM towards CD3ε is at least 100 times greater thanthe K_(d) of the AB when not coupled to the MM towards CD3ε.

In some embodiments, the coupling of the MM reduces the ability of theAB to bind CD3ε such that the dissociation constant (K_(d)) of the ABwhen coupled to the MM towards CD3ε is at least 1000 times greater thanthe K_(d) of the AB when not coupled to the MM towards CD3ε.

In some embodiments, the coupling of the MM reduces the ability of theAB to bind CD3ε such that the dissociation constant (K_(d)) of the ABwhen coupled to the MM towards CD3ε is at least 10,000 times greaterthan the K_(d) of the AB when not coupled to the MM towards CD3ε.

In some embodiments, in the presence of CD3ε, the MM reduces the abilityof the AB to bind CD3ε by at least 90% when the CM is uncleaved, ascompared to when the CM is cleaved when assayed in vitro using a targetdisplacement assay such as, for example, the assay described in PCTPublication No. WO 2010/081173, the contents of which are herebyincorporated by reference in their entirety.

In some embodiments, the MM does not interfere or compete with the AB ofthe activatable antibody in a cleaved state for binding to the CD3εtarget.

In some embodiments, the MM is an amino acid sequence selected from thegroup of those listed in Tables 7 or 8.

In some embodiments, the protease is produced by a tumor that is inproximity to cells that express CD3ε and/or produced by a tumor that isco-localized with CD3ε in a tissue, and the protease cleaves the CM inthe activatable antibody when the activatable antibody is exposed to theprotease.

In some embodiments, the CM is positioned in the activatable antibodysuch that when the activatable antibody is in the uncleaved state,binding of the activatable antibody to CD3ε is reduced to occur with adissociation constant that is at least 20-fold greater than thedissociation constant of an unmodified AB binding to CD3ε, and whereaswhen the activatable antibody is in the cleaved state, the AB bindsCD3ε.

In some embodiments, the CM is positioned in the activatable antibodysuch that when the activatable antibody is in the uncleaved state,binding of the activatable antibody to CD3ε is reduced to occur with adissociation constant that is at least 40-fold greater than thedissociation constant of an unmodified AB binding to CD3ε, and whereaswhen the activatable antibody is in the cleaved state, the AB bindsCD3ε.

In some embodiments, the CM is positioned in the activatable antibodysuch when the activatable antibody is that in the uncleaved state,binding of the activatable antibody to CD3ε is reduced to occur with adissociation constant that is at least 50-fold greater than thedissociation constant of an unmodified AB binding to CD3ε, and whereaswhen the activatable antibody is in the cleaved state, the AB bindsCD3ε.

In some embodiments, the CM is positioned in the activatable antibodysuch that when the activatable antibody is in the uncleaved state,binding of the activatable antibody to CD3ε is reduced to occur with adissociation constant that is at least 100-fold greater than thedissociation constant of an unmodified AB binding to CD3ε, and whereaswhen the activatable antibody is in the cleaved state, the AB bindsCD3ε.

In some embodiments, the CM is positioned in the activatable antibodysuch that when the activatable antibody is in the uncleaved state,binding of the activatable antibody to CD3ε is reduced to occur with adissociation constant that is at least 200-fold greater than thedissociation constant of an unmodified AB binding to CD3ε, and whereaswhen the activatable antibody is in the cleaved state, the AB bindsCD3ε.

In some embodiments, the CM is a polypeptide of up to 15 amino acids inlength.

In some embodiments, the CM includes the amino acid sequence LSGRSDNH(SEQ ID NO: 67). In some embodiments, the cleavable moiety is selectedfor use with a specific protease, for example a protease that is knownto be produced by a tumor that is in proximity to cells that express thetarget of the activatable antibody, e.g., CD3ε, and/or produced by atumor that is co-localized with the target of the activatable antibody.For example, suitable cleavable moieties for use in the activatableanti-CD3ε antibodies of the disclosure are cleaved by at least aprotease such as urokinase, legumain, and/or matriptase (also referredto herein as MT-SP1 or MTSP1). In some embodiments, a suitable cleavablemoiety includes at least one of the following sequences: TGRGPSWV (SEQID NO: 68); SARGPSRW (SEQ ID NO: 69); TARGPSFK (SEQ ID NO: 70); LSGRSDNH(SEQ ID NO: 67); GGWHTGRN (SEQ ID NO: 71); HTGRSGAL (SEQ ID NO: 72);PLTGRSGG (SEQ ID NO: 73); AARGPAIH (SEQ ID NO: 74); RGPAFNPM (SEQ ID NO:75); SSRGPAYL (SEQ ID NO: 76); RGPATPIM (SEQ ID NO: 77); RGPA (SEQ IDNO: 78); GGQPSGMWGW (SEQ ID NO: 79); FPRPLGITGL (SEQ ID NO: 80);VHMPLGFLGP (SEQ ID NO: 81); SPLTGRSG (SEQ ID NO: 82); SAGFSLPA (SEQ IDNO: 83); LAPLGLQRR (SEQ ID NO: 84); SGGPLGVR (SEQ ID NO: 85); and/orPLGL (SEQ ID NO: 86).

In some embodiments, the CM comprises the amino acid sequence LSGRSDNH(SEQ ID NO: 67). In some embodiments, the CM comprises the amino acidsequence TGRGPSWV (SEQ ID NO: 68). In some embodiments, the CM comprisesthe amino acid sequence SARGPSRW (SEQ ID NO: 69). In some embodiments,the CM comprises the amino acid sequence TARGPSFK (SEQ ID NO: 70). Insome embodiments, the CM comprises the amino acid sequence LSGRSDNH (SEQID NO: 67). In some embodiments, the CM comprises the amino acidsequence GGWHTGRN (SEQ ID NO: 71). In some embodiments, the CM comprisesthe amino acid sequence HTGRSGAL (SEQ ID NO: 72). In some embodiments,the CM comprises the amino acid sequence PLTGRSGG (SEQ ID NO: 73). Insome embodiments, the CM comprises the amino acid sequence AARGPAIH (SEQID NO: 74). In some embodiments, the CM comprises the amino acidsequence RGPAFNPM (SEQ ID NO: 75). In some embodiments, the CM comprisesthe amino acid sequence SSRGPAYL (SEQ ID NO: 76). In some embodiments,the CM comprises the amino acid sequence RGPATPIM (SEQ ID NO: 77). Insome embodiments, the CM comprises the amino acid sequence RGPA (SEQ IDNO: 78). In some embodiments, the CM comprises the amino acid sequenceGGQPSGMWGW (SEQ ID NO: 79). In some embodiments, the CM comprises theamino acid sequence FPRPLGITGL (SEQ ID NO: 80). In some embodiments, theCM comprises the amino acid sequence VHMPLGFLGP (SEQ ID NO: 81). In someembodiments, the CM comprises the amino acid sequence SPLTGRSG (SEQ IDNO: 82). In some embodiments, the CM comprises the amino acid sequenceSAGFSLPA (SEQ ID NO: 83). In some embodiments, the CM comprises theamino acid sequence LAPLGLQRR (SEQ ID NO: 84). In some embodiments, theCM comprises the amino acid sequence SGGPLGVR (SEQ ID NO: 85). In someembodiments, the CM comprises the amino acid sequence PLGL (SEQ ID NO:86).

In some embodiments, the CM is a substrate for an MMP and includes thesequence ISSGLLSS (SEQ ID NO: 321); QNQALRMA (SEQ ID NO: 322); AQNLLGMV(SEQ ID NO: 323); STFPFGMF (SEQ ID NO: 324); PVGYTSSL (SEQ ID NO: 325);DWLYWPGI (SEQ ID NO: 326); MIAPVAYR (SEQ ID NO: 327); RPSPMWAY (SEQ IDNO: 328); WATPRPMR (SEQ ID NO: 329); FRLLDWQW (SEQ ID NO: 330); LKAAPRWA(SEQ ID NO: 331); GPSHLVLT (SEQ ID NO: 332); LPGGLSPW (SEQ ID NO: 333);MGLFSEAG (SEQ ID NO: 334); SPLPLRVP (SEQ ID NO: 335); RMHLRSLG (SEQ IDNO: 336); LAAPLGLL (SEQ ID NO: 337); AVGLLAPP (SEQ ID NO: 338); LLAPSHRA(SEQ ID NO: 339); PAGLWLDP (SEQ ID NO: 340); and/or ISSGLSS (SEQ ID NO:341).

In some embodiments, the CM comprises the amino acid sequence ISSGLLSS(SEQ ID NO: 321). In some embodiments, the CM comprises the amino acidsequence QNQALRMA (SEQ ID NO: 322). In some embodiments, the CMcomprises the amino acid sequence AQNLLGMV (SEQ ID NO: 323). In someembodiments, the CM comprises the amino acid sequence STFPFGMF (SEQ IDNO: 324). In some embodiments, the CM comprises the amino acid sequencePVGYTSSL (SEQ ID NO: 325). In some embodiments, the CM comprises theamino acid sequence DWLYWPGI (SEQ ID NO: 326). In some embodiments, theCM comprises the amino acid sequence MIAPVAYR (SEQ ID NO: 327). In someembodiments, the CM comprises the amino acid sequence RPSPMWAY (SEQ IDNO: 328). In some embodiments, the CM comprises the amino acid sequenceWATPRPMR (SEQ ID NO: 329). In some embodiments, the CM comprises theamino acid sequence FRLLDWQW (SEQ ID NO: 330). In some embodiments, theCM comprises the amino acid sequence LKAAPRWA (SEQ ID NO: 331). In someembodiments, the CM comprises the amino acid sequence GPSHLVLT (SEQ IDNO: 332). In some embodiments, the CM comprises the amino acid sequenceLPGGLSPW (SEQ ID NO: 333). In some embodiments, the CM comprises theamino acid sequence MGLFSEAG (SEQ ID NO: 334). In some embodiments, theCM comprises the amino acid sequence SPLPLRVP (SEQ ID NO: 335). In someembodiments, the CM comprises the amino acid sequence RMHLRSLG (SEQ IDNO: 336). In some embodiments, the CM comprises the amino acid sequenceLAAPLGLL (SEQ ID NO: 337). In some embodiments, the CM comprises theamino acid sequence AVGLLAPP (SEQ ID NO: 338). In some embodiments, theCM comprises the amino acid sequence LLAPSHRA (SEQ ID NO: 339). In someembodiments, the CM comprises the amino acid sequence PAGLWLDP (SEQ IDNO: 340). In some embodiments, the CM comprises the amino acid sequenceISSGLSS (SEQ ID NO: 341).

In some embodiments, the CM is a substrate for thrombin. In someembodiments, the CM is a substrate for thrombin and includes thesequence GPRSFGL (SEQ ID NO: 896) or GPRSFG (SEQ ID NO: 897). In someembodiments, the CM comprises the amino acid sequence GPRSFGL (SEQ IDNO: 896). In some embodiments, the CM comprises the amino acid sequenceGPRSFG (SEQ ID NO: 897).

In some embodiments, the CM comprises an amino acid sequence selectedfrom the group consisting of NTLSGRSENHSG (SEQ ID NO: 898); NTLSGRSGNHGS(SEQ ID NO: 899); TSTSGRSANPRG (SEQ ID NO: 900); TSGRSANP (SEQ ID NO:901); VAGRSMRP (SEQ ID NO: 902); VVPEGRRS (SEQ ID NO: 903); ILPRSPAF(SEQ ID NO: 904); MVLGRSLL (SEQ ID NO: 905); QGRAITFI (SEQ ID NO: 906);SPRSIMLA (SEQ ID NO: 907); and SMLRSMPL (SEQ ID NO: 908).

In some embodiments, the CM comprises the amino acid sequenceNTLSGRSENHSG (SEQ ID NO: 898). In some embodiments, the CM comprises theamino acid sequence NTLSGRSGNHGS (SEQ ID NO: 899). In some embodiments,the CM comprises the amino acid sequence TSTSGRSANPRG (SEQ ID NO: 900).In some embodiments, the CM comprises the amino acid sequence TSGRSANP(SEQ ID NO: 901). In some embodiments, the CM comprises the amino acidsequence VAGRSMRP (SEQ ID NO: 902). In some embodiments, the CMcomprises the amino acid sequence VVPEGRRS (SEQ ID NO: 903). In someembodiments, the CM comprises the amino acid sequence ILPRSPAF (SEQ IDNO: 904). In some embodiments, the CM comprises the amino acid sequenceMVLGRSLL (SEQ ID NO: 905). In some embodiments, the CM comprises theamino acid sequence QGRAITFI (SEQ ID NO: 906). In some embodiments, theCM comprises the amino acid sequence SPRSIMLA (SEQ ID NO: 907). In someembodiments, the CM comprises the amino acid sequence and SMLRSMPL (SEQID NO: 908).

In some embodiments, the CM is a substrate for a neutrophil elastase. Insome embodiments, the CM is a substrate for a serine protease. In someembodiments, the CM is a substrate for uPA. In some embodiments, the CMis a substrate for legumain. In some embodiments, the CM is a substratefor matriptase. In some embodiments, the CM is a substrate for acysteine protease. In some embodiments, the CM is a substrate for acysteine protease, such as a cathepsin.

In some embodiments, the CM is a CM1-CM2 substrate and includes thesequence ISSGLLSGRSDNH (SEQ ID NO: 909); ISSGLLSSGGSGGSLSGRSDNH (SEQ IDNO: 910); AVGLLAPPGGTSTSGRSANPRG (SEQ ID NO: 911);TSTSGRSANPRGGGAVGLLAPP (SEQ ID NO: 912); VHMPLGFLGPGGTSTSGRSANPRG (SEQID NO: 913); TSTSGRSANPRGGGVHMPLGFLGP (SEQ ID NO: 914);AVGLLAPPGGLSGRSDNH (SEQ ID NO: 915); LSGRSDNHGGAVGLLAPP (SEQ ID NO:916); VHMPLGFLGPGGLSGRSDNH (SEQ ID NO: 917); LSGRSDNHGGVHMPLGFLGP (SEQID NO: 918); LSGRSDNHGGSGGSISSGLLSS (SEQ ID NO: 919);LSGRSGNHGGSGGSISSGLLSS (SEQ ID NO: 920); ISSGLLSSGGSGGSLSGRSGNH (SEQ IDNO: 921); LSGRSDNHGGSGGSQNQALRMA (SEQ ID NO: 922);QNQALRMAGGSGGSLSGRSDNH (SEQ ID NO: 923); LSGRSGNHGGSGGSQNQALRMA (SEQ IDNO: 924); QNQALRMAGGSGGSLSGRSGNH (SEQ ID NO: 925) and/or ISSGLLSGRSGNH(SEQ ID NO: 926).

In some embodiments, the CM1-CM2 substrate includes the sequenceISSGLLSGRSDNH (SEQ ID NO: 909). In some embodiments, the CM1-CM2substrate comprises the amino acid sequence ISSGLLSSGGSGGSLSGRSDNH (SEQID NO: 910). In some embodiments, the CM1-CM2 substrate comprises theamino acid sequence AVGLLAPPGGTSTSGRSANPRG (SEQ ID NO: 911). In someembodiments, the CM1-CM2 substrate comprises the amino acid sequenceTSTSGRSANPRGGGAVGLLAPP (SEQ ID NO: 912). In some embodiments, theCM1-CM2 substrate comprises the amino acid sequenceVHMPLGFLGPGGTSTSGRSANPRG (SEQ ID NO: 913). In some embodiments, theCM1-CM2 substrate comprises the amino acid sequenceTSTSGRSANPRGGGVHMPLGFLGP (SEQ ID NO: 914). In some embodiments, theCM1-CM2 substrate comprises the amino acid sequence AVGLLAPPGGLSGRSDNH(SEQ ID NO: 915). In some embodiments, the CM1-CM2 substrate comprisesthe amino acid sequence LSGRSDNHGGAVGLLAPP (SEQ ID NO: 916). In someembodiments, the CM1-CM2 substrate comprises the amino acid sequenceVHMPLGFLGPGGLSGRSDNH (SEQ ID NO: 917). In some embodiments, the CM1-CM2substrate comprises the amino acid sequence LSGRSDNHGGVHMPLGFLGP (SEQ IDNO: 918). In some embodiments, the CM1-CM2 substrate comprises the aminoacid sequence LSGRSDNHGGSGGSISSGLLSS (SEQ ID NO: 919). In someembodiments, the CM1-CM2 substrate comprises the amino acid sequenceLSGRSGNHGGSGGSISSGLLSS (SEQ ID NO: 920). In some embodiments, theCM1-CM2 substrate comprises the amino acid sequenceISSGLLSSGGSGGSLSGRSGNH (SEQ ID NO: 921). In some embodiments, theCM1-CM2 substrate comprises the amino acid sequenceLSGRSDNHGGSGGSQNQALRMA (SEQ ID NO: 922). In some embodiments, theCM1-CM2 substrate comprises the amino acid sequenceQNQALRMAGGSGGSLSGRSDNH (SEQ ID NO: 923). In some embodiments, theCM1-CM2 substrate comprises the amino acid sequenceLSGRSGNHGGSGGSQNQALRMA (SEQ ID NO: 924). In some embodiments, theCM1-CM2 substrate comprises the amino acid sequenceQNQALRMAGGSGGSLSGRSGNH (SEQ ID NO: 925). In some embodiments, theCM1-CM2 substrate comprises the amino acid sequence ISSGLLSGRSGNH (SEQID NO: 926).

In some embodiments, the CM is a substrate for a protease selected fromthe group consisting of those shown in Table 3. In some embodiments, theprotease is selected from the group consisting of uPA, legumain,matriptase, ADAM17, BMP-1, TMPRSS3, TMPRSS4, MMP-9, MMP-12, MMP-13, andMMP-14. In some embodiments, the protease is a cathepsin. In someembodiments, the CM is a substrate for a protease selected from thegroup consisting of uPA (urokinase plasminogen activator), legumain andmatriptase. In some embodiments, the protease comprises uPA. In someembodiments, the protease comprises legumain. In some embodiments, theprotease comprises matriptase. In some embodiments, the proteasecomprises a matrix metalloproteinase (MMP).

In some embodiments, the CM is a substrate for at least two proteases.In some embodiments, each protease is selected from the group consistingof those shown in Table 3. In some embodiments, the CM is a substratefor at least two proteases, wherein one of the proteases is selectedfrom the group consisting of uPA, legumain and matriptase and the otherprotease is selected from the group consisting of those shown in Table3. In some embodiments, the CM is a substrate for at least two proteasesselected from the group consisting of uPA, legumain and matriptase.

In some embodiments, the activatable antibody includes at least a firstCM and a second CM. In some embodiments, the first CM and the second CMare each polypeptides of no more than 15 amino acids long. In someembodiments, the first CM and the second CM in the activatable antibodyin the uncleaved state have the structural arrangement from N-terminusto C-terminus as follows: MM-CM1-CM2-AB or AB-CM2-CM1-MM. In someembodiments, at least one of the first CM and the second CM is apolypeptide that functions as a substrate for a protease selected fromthe group consisting of uPA, legumain, and matriptase. In someembodiments, the first CM is cleaved by a first cleaving agent selectedfrom the group consisting of uPA, legumain, and matriptase in a targettissue and the second CM is cleaved by a second cleaving agent in atarget tissue. In some embodiments, the other protease is selected fromthe group consisting of those shown in Table 3. In some embodiments, thefirst cleaving agent and the second cleaving agent are the same proteaseselected from the group consisting of uPA, legumain, and matriptase, andthe first CM and the second CM are different substrates for theprotease. In some embodiments, the first cleaving agent and the secondcleaving agent are the same protease selected from the group consistingof those shown in Table 3. In some embodiments, the first cleaving agentand the second cleaving agent are different proteases. In someembodiments, the first cleaving agent and the second cleaving agent areproduced by a tumor that is in proximity to cells that express thetarget and/or produced by a tumor that is co-localized in the targettissue. In some embodiments, the first CM and the second CM are cleavedby at least one cleaving agent in the target tissue.

In some embodiments, the activatable antibody is exposed to and cleavedby a protease such that, when the activatable antibody is in theactivated or cleaved state, the activated antibody includes a lightchain amino acid sequence that includes at least a portion of LP2 and/orCM sequence after the protease has cleaved the CM.

In some embodiments, the MM and the CM include an amino acid sequenceselected from the group consisting of those presented herein.

In some embodiments, the activatable antibody also includes a signalpeptide. In some embodiments, the signal peptide is conjugated to theactivatable antibody via a spacer. In some embodiments, the spacer isconjugated to the activatable antibody in the absence of a signalpeptide. In some embodiments, the spacer is joined directly to the MM ofthe activatable antibody.

In some embodiments, the activatable antibody in an uncleaved statecomprises a spacer that is joined directly to the MM and has thestructural arrangement from N-terminus to C-terminus of spacer-MM-CM-AB.In some embodiments, the spacer includes at least the amino acidsequence QGQSGQG (SEQ ID NO: 407). In some embodiments, the spacerincludes at least the amino acid sequence QGQSGQ (SEQ ID NO: 87). Insome embodiments, the spacer includes at least the amino acid sequenceQGQSG (SEQ ID NO: 408). In some embodiments, the spacer includes atleast the amino acid sequence QGQS (SEQ ID NO: 409), In someembodiments, the spacer includes at least the amino acid sequence QGQ(SEQ ID NO: 410). In some embodiments, the spacer includes at least theamino acid sequence QG (SEQ ID NO: 411). In some embodiments, the spacerincludes at least the amino acid residue Q. In some embodiments, the MMand spacer comprise an amino acid sequence selected from the sequenceslisted in Table 7 or Table 8.

In some embodiments, the activatable antibody includes a heavy chainsequence selected from the group consisting of SEQ ID NOs: 446, 452,454, 456, 460, 462, 464, 466, 470, 472, 476, 478, 480, 486, 488, 490,492, 494, 496, 498, 500, 502, 504, 510, 512, 514, 518, 524, 526, 530,532, 534, 536, 538, 540, 542, 544, and 546.

In some embodiments, the activatable antibody includes a light chainsequence selected from the group consisting of SEQ ID NOs: 448, 450,458, 468, 474, 482, 484, 508, 516, and 520.

In some embodiments, the activatable antibody includes a heavy chainsequence selected from the group consisting of SEQ ID NOs: 446, 452,454, 456, 460, 462, 464, 466, 470, 472, 476, 478, 480, 486, 488, 490,492, 494, 496, 498, 500, 502, 504, 510, 512, 514, 518, 524, 526, 530,532, 534, 536, 538, 540, 542, 544, and 546; and a light chain sequenceselected from the group consisting of SEQ ID NOs: 448, 450, 458, 468,474, 482, 484, 508, 516, and 520.

In some embodiments, the activatable antibody includes a heavy chainsequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or99% identical to an amino acid sequence selected from the groupconsisting of SEQ ID NOs: 446, 452, 454, 456, 460, 462, 464, 466, 470,472, 476, 478, 480, 486, 488, 490, 492, 494, 496, 498, 500, 502, 504,510, 512, 514, 518, 524, 526, 530, 532, 534, 536, 538, 540, 542, 544,and 546.

In some embodiments, the activatable antibody includes a light chainsequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or99% identical to an amino acid sequence selected from the groupconsisting of SEQ ID NOs: 448, 450, 458, 468, 474, 482, 484, 508, 516,and 520.

In some embodiments, the activatable antibody includes a heavy chainsequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or99% identical to an amino acid sequence selected from the groupconsisting of SEQ ID NOs: 446, 452, 454, 456, 460, 462, 464, 466, 470,472, 476, 478, 480, 486, 488, 490, 492, 494, 496, 498, 500, 502, 504,510, 512, 514, 518, 524, 526, 530, 532, 534, 536, 538, 540, 542, 544,and 546; and a light chain sequence that is at least 90%, 91%, 92%, 93%,94%, 95%, 96%, 97%, 98% or 99% identical to an amino acid sequenceselected from the group consisting of SEQ ID NOs: 448, 450, 458, 468,474, 482, 484, 508, 516, and 520.

In some embodiments, the activatable antibody includes the amino acidsequence of SEQ ID NO: 506. In some embodiments, the activatableantibody includes an amino acid sequence that is at least 90%, 91%, 92%,93%, 94%, 95%, 96%, 97%, 98% or 99% identical to the amino acid sequenceof SEQ ID NO: 506.

In some embodiments, the activatable antibody includes the amino acidsequence of SEQ ID NO: 587 or SEQ ID NO: 588. In some embodiments, theactivatable antibody includes an amino acid sequence that is at least90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identical to theamino acid sequence of SEQ ID NO: 587 or SEQ ID NO: 588.

In some embodiments, the activatable antibody also includes an agentconjugated to the AB. In some embodiments, the agent is a therapeuticagent. In some embodiments, the agent is a detectable moiety. In someembodiments, the detectable moiety is a diagnostic agent. In someembodiments, the agent is conjugated to the AB via a linker. In someembodiments, the linker is a cleavable linker. In some embodiments, thelinker is a non-cleavable linker.

In some embodiments, the activatable antibody also includes a detectablemoiety. In some embodiments, the detectable moiety is a diagnosticagent.

In some embodiments, the AB of the activatable antibody naturallycontains one or more disulfide bonds. In some embodiments, the AB can beengineered to include one or more disulfide bonds.

In some embodiments, the serum half-life of the multispecificactivatable antibody is longer than that of the correspondingmultispecific antibody; e.g., the pK of the multispecific activatableantibody is longer than that of the corresponding multispecificantibody. In some embodiments, the serum half-life of the multispecificactivatable antibody is similar to that of the correspondingmultispecific antibody. In some embodiments, the serum half-life of themultispecific activatable antibody is at least 15 days when administeredto an organism. In some embodiments, the serum half-life of themultispecific activatable antibody is at least 12 days when administeredto an organism. In some embodiments, the serum half-life of themultispecific activatable antibody is at least 11 days when administeredto an organism. In some embodiments, the serum half-life of themultispecific activatable antibody is at least 10 days when administeredto an organism. In some embodiments, the serum half-life of themultispecific activatable antibody is at least 9 days when administeredto an organism. In some embodiments, the serum half-life of themultispecific activatable antibody is at least 8 days when administeredto an organism. In some embodiments, the serum half-life of themultispecific activatable antibody is at least 7 days when administeredto an organism. In some embodiments, the serum half-life of themultispecific activatable antibody is at least 6 days when administeredto an organism. In some embodiments, the serum half-life of theactivatable antibody is at least 5 days when administered to anorganism. In some embodiments, the serum half-life of the activatableantibody is at least 4 days when administered to an organism. In someembodiments, the serum half-life of the activatable antibody is at least3 days when administered to an organism. In some embodiments, the serumhalf-life of the activatable antibody is at least 2 days whenadministered to an organism. In some embodiments, the serum half-life ofthe activatable antibody is at least 24 hours when administered to anorganism. In some embodiments, the serum half-life of the activatableantibody is at least 20 hours when administered to an organism. In someembodiments, the serum half-life of the activatable antibody is at least18 hours when administered to an organism. In some embodiments, theserum half-life of the activatable antibody is at least 16 hours whenadministered to an organism. In some embodiments, the serum half-life ofthe activatable antibody is at least 14 hours when administered to anorganism. In some embodiments, the serum half-life of the activatableantibody is at least 12 hours when administered to an organism. In someembodiments, the serum half-life of the activatable antibody is at least10 hours when administered to an organism. In some embodiments, theserum half-life of the activatable antibody is at least 8 hours whenadministered to an organism. In some embodiments, the serum half-life ofthe activatable antibody is at least 6 hours when administered to anorganism. In some embodiments, the serum half-life of the activatableantibody is at least 4 hours when administered to an organism. In someembodiments, the serum half-life of the activatable antibody is at least3 hours when administered to an organism.

In some embodiments, the activatable anti-CD3ε antibody is monospecific.In some embodiments, the activatable anti-CD3ε antibody ismultispecific, e.g., by way of non-limiting example, bispecific ortrifunctional. In some embodiments, the activatable anti-CD3ε antibodyis formulated as part of a pro-Bispecific T Cell Engager (BITE)molecule. In some embodiments, the activatable anti-CD3ε antibody isformulated as part of a pro-Chimeric Antigen Receptor (CAR) modified Tcell or other engineered receptor.

The compositions and methods provided herein enable the attachment ofone or more agents to one or more cysteine residues in the AB withoutcompromising the activity (e.g., the masking, activating or bindingactivity) of the activatable anti-CD3ε antibody. In some embodiments,the compositions and methods provided herein enable the attachment ofone or more agents to one or more cysteine residues in the AB withoutreducing or otherwise disturbing one or more disulfide bonds within theMM. The compositions and methods provided herein produce an activatableanti-CD3ε antibody that is conjugated to one or more agents, e.g., anyof a variety of therapeutic, diagnostic and/or prophylactic agents,preferably without any of the agent(s) being conjugated to the MM of theactivatable anti-CD3ε antibody. The compositions and methods providedherein produce conjugated activatable anti-CD3ε antibodies in which theMM retains the ability to effectively and efficiently mask the AB of theactivatable antibody in an uncleaved state. The compositions and methodsprovided herein produce conjugated activatable anti-CD3ε antibodies inwhich the activatable antibody is still activated, i.e., cleaved, in thepresence of a protease that can cleave the CM.

In some embodiments, the anti-CD3ε antibodies and/or the activatableanti-CD3ε antibodies described herein are used in conjunction with oneor more additional agents or a combination of additional agents.Suitable additional agents include current pharmaceutical and/orsurgical therapies for an intended application, such as, for example,cancer. For example, the anti-CD3ε antibodies and/or activatableanti-CD3ε antibodies can be used in conjunction with an additionalchemotherapeutic or anti-neoplastic agent.

In some embodiments, the anti-CD3ε antibody and/or activatable anti-CD3εantibody and additional agent are formulated into a single therapeuticcomposition, and the anti-CD3ε antibody and/or activatable anti-CD3εantibody and additional agent are administered simultaneously. In someembodiments, the anti-CD3ε antibody and/or activatable anti-CD3εantibody and additional agent are separate from each other, e.g., eachis formulated into a separate therapeutic composition, and the anti-CD3εantibody and/or activatable anti-CD3ε antibody and the additional agentare administered simultaneously, or the anti-CD3ε antibody and/oractivatable anti-CD3ε antibody and the additional agent are administeredat different times during a treatment regimen. For example, theanti-CD3ε antibody and/or activatable anti-CD3ε antibody is administeredprior to the administration of the additional agent, the anti-CD3εantibody and/or activatable anti-CD3ε antibody is administeredsubsequent to the administration of the additional agent, or theanti-CD3ε antibody and/or activatable anti-CD3ε antibody and theadditional agent are administered in an alternating fashion. Asdescribed herein, the anti-CD3ε antibody and/or activatable anti-CD3εantibody and additional agent are administered in single doses or inmultiple doses.

The disclosure also provides an isolated nucleic acid molecule encodingat least a portion of an activatable anti-CD3ε antibody describedherein, as well as vectors that include these isolated nucleic acidsequences and/or one or more nucleic acid molecules encoding ananti-CD3ε antibody described herein, such as for example, at least afirst nucleic acid encoding at least a portion of the heavy chain of theactivatable antibody and a second nucleic acid encoding at least aportion of the light chain of the activatable antibody. The disclosureprovides methods of producing an activatable antibody by culturing acell under conditions that lead to expression of the activatableantibody, wherein the cell comprises such a nucleic acid molecule(s). Insome embodiments, the cell comprises such a vector.

In some embodiments, the activatable antibody is encoded by a heavychain nucleic acid sequence selected from the group consisting of SEQ IDNOs: 445, 451, 453, 455, 459, 461, 463, 465, 469, 471, 475, 477, 479,485, 487, 489, 491, 493, 495, 497, 499, 501, 503, 511, 513, 517, 522,525, 529, 531, 533, 535, 537, 539, 541, 543, and 545.

In some embodiments, the activatable antibody is encoded by a lightchain nucleic acid selected from the group consisting of SEQ ID NOs:447, 449, 457, 467, 473, 481, 483, 507, 515, and 519.

In some embodiments, the activatable antibody is encoded by a heavychain nucleic acid sequence selected from the group consisting of SEQ IDNOs: 445, 451, 453, 455, 459, 461, 463, 465, 469, 471, 475, 477, 479,485, 487, 489, 491, 493, 495, 497, 499, 501, 503, 511, 513, 517, 522,525, 529, 531, 533, 535, 537, 539, 541, 543, and 545; and a light chainnucleic acid sequence selected from the group consisting of SEQ ID NOs:447, 449, 457, 467, 473, 481, 483, 507, 515, and 519.

In some embodiments, the activatable antibody is encoded by a heavychain nucleic acid sequence that is at least 90%, 91%, 92%, 93%, 94%,95%, 96%, 97%, 98% or 99% identical to a sequence selected from thegroup consisting of SEQ ID NOs: 445, 451, 453, 455, 459, 461, 463, 465,469, 471, 475, 477, 479, 485, 487, 489, 491, 493, 495, 497, 499, 501,503, 511, 513, 517, 522, 525, 529, 531, 533, 535, 537, 539, 541, 543,and 545.

In some embodiments, the activatable antibody is encoded by a lightnucleic acid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%,96%, 97%, 98% or 99% identical to a sequence selected from the groupconsisting of SEQ ID NOs: 447, 449, 457, 467, 473, 481, 483, 507, 515,and 519.

In some embodiments, the activatable antibody is encoded by a heavychain nucleic acid sequence that is at least 90%, 91%, 92%, 93%, 94%,95%, 96%, 97%, 98% or 99% identical to a sequence selected from thegroup consisting of SEQ ID NOs: 445, 451, 453, 455, 459, 461, 463, 465,469, 471, 475, 477, 479, 485, 487, 489, 491, 493, 495, 497, 499, 501,503, 511, 513, 517, 522, 525, 529, 531, 533, 535, 537, 539, 541, 543,and 545; and a light chain sequence that is at least 90%, 91%, 92%, 93%,94%, 95%, 96%, 97%, 98% or 99% identical to a sequence selected from thegroup consisting of SEQ ID NOs: 447, 449, 457, 467, 473, 481, 483, 507,515, and 519.

In some embodiments, the activatable antibody is encoded by a nucleicacid that includes the nucleic acid sequence of SEQ ID NO: 505. In someembodiments, the activatable antibody includes a nucleic acid sequencethat is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99%identical to the sequence of SEQ ID NO: 505.

The disclosure also provides a method of manufacturing activatableantibodies that binds CD3ε in an activated state by (a) culturing a cellcomprising a nucleic acid construct that encodes the activatableantibody under conditions that lead to expression of the activatableantibody, wherein the activatable antibody comprises a masking moiety(MM), a cleavable moiety (CM), and an antibody or an antigen bindingfragment thereof (AB) that specifically binds CD3ε, and (b) recoveringthe activatable antibody.

The disclosure also provides a method of manufacturing activatableantibodies that binds CD3ε in an activated state by (a) culturing a cellcomprising a nucleic acid construct that encodes the activatableantibody under conditions that lead to expression of the activatableantibody, wherein the activatable antibody comprises a masking moiety(MM), a cleavable moiety (CM), and an antibody or an antigen bindingfragment thereof (AB) that specifically binds CD3ε, (i) wherein the CMis a polypeptide that includes an amino acid sequence that functions asa substrate for a protease; and (ii) wherein the CM is positioned in theactivatable antibody such that, when the activatable antibody is in anuncleaved state, the MM interferes with specific binding of the AB toCD3 and when the activatable antibody is in a cleaved state the MM doesnot interfere or compete with specific binding of the AB to CD3ε; and(b) recovering the activatable antibody.

The disclosure also provides multispecific antibodies that bind theepsilon chain of CD3 (CD3ε) and a second target, wherein the antibodycomprises a first antibody or antigen-binding fragment thereof (AB1)that binds the epsilon chain of CD3 (CD3ε) and a second antibody orantigen-binding fragment thereof (AB2) that binds a second target, andwherein AB1 comprises a VH CDR1 sequence that includes an amino acidsequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%,99% or more identical to the amino acid sequence TYAMN (SEQ ID NO: 53);a VH CD2 sequence that includes an amino acid sequence that is at least90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical tothe amino acid sequence RIRSKYNNYATYYADSVKD (SEQ ID NO: 54); a VH CDR3sequence that includes an amino acid sequence that is at least 90%, 91%,92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical to the aminoacid sequence HGNFGNSYVSWFAY (SEQ ID NO: 55), and a combination of a VLCDR1 sequence, a VL CDR2 sequence, and a VL CDR3 sequence, wherein atleast one of the VL CDR1 sequence, the VL CDR2 sequence, and the VL CDR3sequence is selected from a VL CDR1 sequence that includes an amino acidsequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%,99% or more identical to the amino acid sequence RSSTGAVTTSNYAN (SEQ IDNO: 56); a VL CDR2 sequence that includes an amino acid sequence that isat least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or moreidentical to the amino acid sequence GTNKRAP (SEQ ID NO: 57); and a VLCDR3 sequence that includes an amino acid sequence that is at least 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical to theamino acid sequence ALWYSNLWV (SEQ ID NO: 58).

In some embodiments, the multispecific anti-CD3ε antibody orantigen-binding fragment thereof includes a VH CDR1 sequence thatincludes at least the amino acid sequence TYAMN (SEQ ID NO: 53); a VHCD2 sequence that includes at least the amino acid sequenceRIRSKYNNYATYYADSVKD (SEQ ID NO: 54); a VH CDR3 sequence that includes atleast the amino acid sequence HGNFGNSYVSWFAY (SEQ ID NO: 55), a VL CDR1sequence that includes at least the amino acid sequence RSSTGAVTTSNYAN(SEQ ID NO: 56); a VL CDR2 sequence that includes at least the aminoacid sequence GTNKRAP (SEQ ID NO: 57); and a VL CDR3 sequence thatincludes at least the amino acid sequence ALWYSNLWV (SEQ ID NO: 58).

In some embodiments, AB1 comprises a variable heavy chain (Hv)comprising the amino acid sequence of SEQ ID NO: 4 and a variable lightchain (Lv) comprising the amino acid sequence of SEQ ID NO: 2.

In some embodiments, AB1 comprises a variable heavy chain (Hv)comprising an amino acid sequence that is at least 90%, 91%, 92%, 93%,94%, 95%, 96%, 97%, 98%, 99% or more identical to the amino acidsequence of SEQ ID NO: 4 and a variable light chain (Lv) comprising anamino acid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%,97%, 98%, 99% or more identical to the amino acid sequence of SEQ ID NO:2.

In some embodiments, AB1 comprises a scFv fragment. In some embodiments,the scFv fragment comprises the amino acid sequence of SEQ ID NO: 6. Insome embodiments, the scFv fragment comprises the amino acid sequence ofSEQ ID NO: 30. In some embodiments, the scFv fragment comprises an aminoacid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%,98%, 99% or more identical to the amino acid sequence of SEQ ID NO: 6.In some embodiments, the scFv fragment comprises an amino acid sequencethat is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% ormore identical to the amino acid sequence of SEQ ID NO: 30.

In some embodiments, the multispecific antibody includes an agentconjugated to the antibody. In some embodiments, the agent is atherapeutic agent, a detectable moiety or a diagnostic agent. In someembodiments, the agent is conjugated to the antibody via a linker. Insome embodiments, the linker is a cleavable linker. In some embodiments,the linker is a non-cleavable linker. In some embodiments, the linker isa non-cleavable linker.

The present disclosure also provides multispecific antibodies andmultispecific activatable antibodies that bind at least CD3ε. Themultispecific antibodies provided herein are antibodies that recognizetwo or more different antigens or epitopes, where at least one antigenor epitope is CD3ε. The multispecific activatable antibodies providedherein are multispecific antibodies that include at least one maskingmoiety (MM) linked to at least one antigen- or epitope-binding domain ofthe multispecific antibody such that coupling of the MM reduces theability of the antigen- or epitope-binding domain to bind its target. Insome embodiments, the MM is coupled to the antigen- or epitope-bindingdomain of the multispecific antibody via a cleavable moiety (CM) thatfunctions as a substrate for a protease. The activatable multispecificantibodies provided herein are stable in circulation, activated atintended sites of therapy and/or diagnosis but not in normal, i.e.,healthy tissue, and, when activated, exhibit binding to a target that isat least comparable to the corresponding, unmodified multispecificantibody.

In some embodiments, the multispecific antibodies and/or multispecificactivatable antibodies are designed to engage immune effector cells,also referred to herein as immune effector cell engaging multispecificantibodies and/or immune effector cell engaging multispecificactivatable antibodies. In some embodiments, the multispecificantibodies and/or multispecific activatable antibodies are designed toengage leukocytes, also referred to herein as leukocyte engagingmultispecific antibodies and/or leukocyte engaging multispecificactivatable antibodies. In some embodiments, the multispecificantibodies and/or multispecific activatable antibodies are designed toengage T cells, also referred to herein as T-cell engaging multispecificantibodies and/or T-cell engaging multispecific activatable antibodies.In some embodiments, the multispecific antibodies and/or multispecificactivatable antibodies engage a surface antigen on a leukocyte, such ason a T cell, on a natural killer (NK) cell, on a myeloid mononuclearcell, on a macrophage, and/or on another immune effector cell. In someembodiments, the immune effector cell is a leukocyte. In someembodiments, the immune effector cell is a T cell. In some embodiments,the immune effector cell is a NK cell. In some embodiments, the immuneeffector cell is a mononuclear cell, such as a myeloid mononuclear cell.In some embodiments, the T-cell engaging multispecific antibodies and/orT-cell engaging multispecific activatable antibodies bind CD3ε.

In some embodiments, the immune effector cell engaging multispecificantibodies include a targeting antibody or antigen-binding fragmentthereof and an immune effector cell engaging antibody or antigen-bindingportion thereof that includes an anti-CD3ε antibody or antigen-bindingfragment thereof. In some embodiments, the immune effector cell engagingmultispecific antibodies include a cancer targeting antibody orantigen-binding fragment thereof and an immune effector cell engagingantibody or antigen-binding portion thereof. In some embodiments, theimmune effector cell engaging multispecific antibodies include a cancertargeting IgG antibody or antigen-binding fragment thereof and an immuneeffector cell engaging scFv. In some embodiments, the immune effectorcell is a leukocyte. In some embodiments, the immune effector cell is aT cell. In some embodiments, the immune effector cell is a NK cell. Insome embodiments, the immune effector cell is a myeloid mononuclearcell.

In some embodiments, the T-cell engaging multispecific antibody includesan anti-CD3ε antibody or antigen-binding fragment thereof and atargeting antibody or antigen-binding fragment thereof. In someembodiments, the T-cell engaging multispecific antibody includes ananti-CD3ε scFv and a targeting antibody or antigen-binding fragmentthereof. In some embodiments, the T-cell engaging multispecific antibodyincludes an anti-CD3ε antibody or antigen-binding fragment thereof and acancer-targeting antibody or antigen-binding fragment thereof. In someembodiments, the T-cell engaging multispecific antibody includes ananti-CD3ε scFv and a cancer targeting antibody or antigen-bindingfragment thereof. In some embodiments, the T-cell engaging multispecificantibody includes an anti-CD3ε antibody or antigen-binding fragmentthereof and a cancer-targeting IgG antibody or antigen-binding fragmentthereof. In some embodiments, the T-cell engaging multispecific antibodyincludes an anti-CD3ε scFv and a cancer targeting IgG antibody orantigen-binding fragment thereof. In some embodiments, the T-cellengaging multispecific antibody includes an anti-CD3 epsilon (CD3ε) scFvthat is derived from OKT3. In some embodiments, the T-cell engagingmultispecific antibody includes an anti-CD3 epsilon (CD3ε) scFv that isderived from SP34 (available from BD Biosciences Cat#556610).

In some embodiments, the T-cell engaging multispecific activatableantibodies include a targeting antibody or antigen-binding fragmentthereof and anti-CD3ε antibody or antigen-binding fragment thereof,where at least one of the targeting antibody or antigen-binding fragmentthereof and/or the anti-CD3ε antibody or antigen-binding portion thereofis masked. In some embodiments, the anti-CD3ε antibody or antigenbinding fragment thereof includes a first antibody or antigen-bindingfragment thereof (AB1) that binds CD3ε, where the AB1 is attached to amasking moiety (MM1) such that coupling of the MM1 reduces the abilityof the AB1 to bind CD3ε. In some embodiments, the targeting antibody orantigen-binding fragment thereof includes a second antibody or fragmentthereof that includes a second antibody or antigen-binding fragmentthereof (AB2) that binds a second target, where the AB2 is attached to amasking moiety (MM2) such that coupling of the MM2 reduces the abilityof the AB2 to bind the second target. In some embodiments, the anti-CD3εantibody or antigen binding fragment thereof includes a first antibodyor antigen-binding fragment thereof (AB1) that binds CD3ε, where the AB1is attached to a masking moiety (MM1) such that coupling of the MM1reduces the ability of the AB1 to bind CD3ε, and the targeting antibodyor antigen-binding fragment thereof includes a second antibody orfragment thereof that includes a second antibody or antigen-bindingfragment thereof (AB2) that binds a second target, where the AB2 isattached to a masking moiety (MM2) such that coupling of the MM2 reducesthe ability of the AB2 to bind the second target.

In some embodiments, the T-cell engaging multispecific activatableantibodies include a cancer targeting antibody or antigen-bindingfragment thereof and an anti-CD3ε antibody or antigen-binding portionthereof, where at least one of the cancer targeting antibody orantigen-binding fragment thereof and/or the anti-CD3ε antibody orantigen-binding portion thereof is masked. In some embodiments, theanti-CD3ε antibody or antigen binding fragment thereof includes a firstantibody or antigen-binding fragment thereof (AB1) that binds CD3ε,where the AB1 is attached to a masking moiety (MM1) such that couplingof the MM1 reduces the ability of the AB1 to bind CD3ε. In someembodiments, the cancer targeting antibody or antigen-binding fragmentthereof includes a second antibody or fragment thereof that includes asecond antibody or antigen-binding fragment thereof (AB2) that binds asecond, cancer-related target, where the AB2 is attached to a maskingmoiety (MM2) such that coupling of the MM2 reduces the ability of theAB2 to bind the second, cancer-related target. In some embodiments, theanti-CD3ε antibody or antigen binding fragment thereof includes a firstantibody or antigen-binding fragment thereof (AB1) that binds CD3ε,where the AB1 is attached to a masking moiety (MM1) such that couplingof the MM1 reduces the ability of the AB1 to bind CD3ε, and the cancertargeting antibody or antigen-binding fragment thereof includes a secondantibody or fragment thereof that includes a second antibody orantigen-binding fragment thereof (AB2) that binds a second,cancer-related target, where the AB2 is attached to a masking moiety(MM2) such that coupling of the MM2 reduces the ability of the AB2 tobind the second, cancer-related target.

In some embodiments, the T-cell engaging multispecific activatableantibodies include a cancer targeting IgG antibody or antigen-bindingfragment thereof and an anti-CD3ε, where at least one of the cancertargeting IgG antibody or antigen-binding fragment thereof and/or theanti-CD3ε antibody or antigen-binding portion thereof is masked. In someembodiments, the anti-CD3ε antibody or antigen binding fragment thereofincludes a first antibody or antigen-binding fragment thereof (AB1) thatbinds CD3ε, where the AB1 is attached to a masking moiety (MM1) suchthat coupling of the MM1 reduces the ability of the AB1 to bind CD3ε. Insome embodiments, the cancer targeting IgG antibody or antigen-bindingfragment thereof includes a second antibody or fragment thereof thatincludes a second antibody or antigen-binding fragment thereof (AB2)that binds a second, cancer-related target, where the AB2 is attached toa masking moiety (MM2) such that coupling of the MM2 reduces the abilityof the AB2 to bind the second, cancer-related target. In someembodiments, the anti-CD3ε antibody or antigen binding fragment thereofincludes a first antibody or antigen-binding fragment thereof (AB1) thatbinds CD3ε, where the AB1 is attached to a masking moiety (MM1) suchthat coupling of the MM1 reduces the ability of the AB1 to bind CD3ε,and the cancer targeting IgG antibody or antigen-binding fragmentthereof includes a second antibody or fragment thereof that includes asecond antibody or antigen-binding fragment thereof (AB2) that binds asecond, cancer-related target, where the AB2 is attached to a maskingmoiety (MM2) such that coupling of the MM2 reduces the ability of theAB2 to bind the second, cancer-related target.

In some embodiments, the T-cell engaging multispecific activatableantibody includes an anti-CD3ε scFv and a targeting antibody orantigen-binding fragment thereof, where at least one of the anti-CD3εscFv and/or the targeting antibody or antigen-binding portion thereof ismasked. In some embodiments, the CD3ε scFv includes a first antibody orantigen-binding fragment thereof (AB1) that binds CD3ε, where the AB1 isattached to a masking moiety (MM1) such that coupling of the MM1 reducesthe ability of the AB1 to bind CD3ε. In some embodiments, the targetingantibody or antigen-binding fragment thereof includes a second antibodyor fragment thereof that includes a second antibody or antigen-bindingfragment thereof (AB2) that binds a second target, where the AB2 isattached to a masking moiety (MM2) such that coupling of the MM2 reducesthe ability of the AB2 to bind the second target. In some embodiments,the CD3ε scFv includes a first antibody or antigen-binding fragmentthereof (AB1) that binds CD3ε, where the AB1 is attached to a maskingmoiety (MM1) such that coupling of the MM1 reduces the ability of theAB1 to bind CD3ε, and the targeting antibody or antigen-binding fragmentthereof includes a second antibody or fragment thereof that includes asecond antibody or antigen-binding fragment thereof (AB2) that binds asecond target, where the AB2 is attached to a masking moiety (MM2) suchthat coupling of the MM2 reduces the ability of the AB2 to bind thesecond target.

In some embodiments, the T-cell engaging multispecific activatableantibody includes an anti-CD3ε scFv and a cancer targeting antibody orantigen-binding fragment thereof, where at least one of the anti-CD3εscFv and/or the cancer targeting antibody or antigen-binding portionthereof is masked. In some embodiments, the CD3ε scFv includes a firstantibody or antigen-binding fragment thereof (AB1) that binds CD3ε,where the AB1 is attached to a masking moiety (MM1) such that couplingof the MM1 reduces the ability of the AB1 to bind CD3ε. In someembodiments, the cancer targeting antibody or antigen-binding fragmentthereof includes a second antibody or fragment thereof that includes asecond antibody or antigen-binding fragment thereof (AB2) that binds asecond, cancer-related target, where the AB2 is attached to a maskingmoiety (MM2) such that coupling of the MM2 reduces the ability of theAB2 to bind the second, cancer-related target. In some embodiments, theCD3ε scFv includes a first antibody or antigen-binding fragment thereof(AB1) that binds CD3ε, where the AB1 is attached to a masking moiety(MM1) such that coupling of the MM1 reduces the ability of the AB1 tobind CD3ε, and the cancer targeting antibody or antigen-binding fragmentthereof includes a second antibody or fragment thereof that includes asecond antibody or antigen-binding fragment thereof (AB2) that binds asecond, cancer-related target, where the AB2 is attached to a maskingmoiety (MM2) such that coupling of the MM2 reduces the ability of theAB2 to bind the second, cancer-related target.

In some embodiments, the T-cell engaging multispecific activatableantibody includes an anti-CD3ε scFv and a cancer targeting IgG antibodyor antigen-binding fragment thereof, where at least one of the anti-CD3εscFv and/or the cancer targeting IgG antibody or antigen-binding portionthereof is masked. In some embodiments, the CD3ε scFv includes a firstantibody or antigen-binding fragment thereof (AB1) that binds CD3ε,where the AB1 is attached to a masking moiety (MM1) such that couplingof the MM1 reduces the ability of the AB1 to bind CD3ε. In someembodiments, the cancer targeting IgG antibody or antigen-bindingfragment thereof includes a second antibody or fragment thereof thatincludes a second antibody or antigen-binding fragment thereof (AB2)that binds a second, cancer-related target, where the AB2 is attached toa masking moiety (MM2) such that coupling of the MM2 reduces the abilityof the AB2 to bind the second, cancer-related target. In someembodiments, the CD3ε scFv includes a first antibody or antigen-bindingfragment thereof (AB1) that binds CD3ε, where the AB1 is attached to amasking moiety (MM1) such that coupling of the MM1 reduces the abilityof the AB1 to bind CD3ε, and the cancer targeting antibody IgG orantigen-binding fragment thereof includes a second antibody or fragmentthereof that includes a second antibody or antigen-binding fragmentthereof (AB2) that binds a second, cancer-related target, where the AB2is attached to a masking moiety (MM2) such that coupling of the MM2reduces the ability of the AB2 to bind the second, cancer-relatedtarget.

In some embodiments, the T-cell engaging multispecific activatableantibody includes an anti-CD3 epsilon (CD3ε) scFv that is derived fromOKT3, where at least one of the targeting antibody or antigen-bindingfragment thereof and/or the OKT3 scFv or OKT3-derived scFv is masked. Insome embodiments, the OKT3 scFv or OKT3-derived scFv includes a firstantibody or antigen-binding fragment thereof (AB1) that binds CD3ε,where the AB1 is attached to a masking moiety (MM1) such that couplingof the MM1 reduces the ability of the AB1 to bind CD3ε. In someembodiments, the targeting antibody or antigen-binding fragment thereofincludes a second antibody or fragment thereof that includes a secondantibody or antigen-binding fragment thereof (AB2) that binds a secondtarget, where the AB2 is attached to a masking moiety (MM2) such thatcoupling of the MM2 reduces the ability of the AB2 to bind the secondtarget. In some embodiments, the OKT3 scFv or OKT3-derived scFv includesa first antibody or antigen-binding fragment thereof (AB1) that bindsCD3ε, where the AB1 is attached to a masking moiety (MM1) such thatcoupling of the MM1 reduces the ability of the AB1 to bind CD3ε, and thetargeting antibody or antigen-binding fragment thereof includes a secondantibody or fragment thereof that includes a second antibody orantigen-binding fragment thereof (AB2) that binds a second target, wherethe AB2 is attached to a masking moiety (MM2) such that coupling of theMM2 reduces the ability of the AB2 to bind the second target.

In some embodiments, the T-cell engaging multispecific activatableantibody includes an OKT3 scFv or OKT3-derived scFv and a cancertargeting antibody or antigen-binding fragment thereof, where at leastone of the OKT3 scFv or OKT3-derived scFv and/or the cancer targetingantibody or antigen-binding portion thereof is masked. In someembodiments, the OKT3 scFv or OKT3-derived scFv includes a firstantibody or antigen-binding fragment thereof (AB1) that binds CD3ε,where the AB1 is attached to a masking moiety (MM1) such that couplingof the MM1 reduces the ability of the AB1 to bind CD3ε. In someembodiments, the cancer targeting antibody or antigen-binding fragmentthereof includes a second antibody or fragment thereof that includes asecond antibody or antigen-binding fragment thereof (AB2) that binds asecond, cancer-related target, where the AB2 is attached to a maskingmoiety (MM2) such that coupling of the MM2 reduces the ability of theAB2 to bind the second, cancer-related target. In some embodiments, theOKT3 scFv or OKT3-derived scFv includes a first antibody orantigen-binding fragment thereof (AB1) that binds CD3ε, where the AB1 isattached to a masking moiety (MM1) such that coupling of the MM1 reducesthe ability of the AB1 to bind CD3ε, and the cancer targeting antibodyor antigen-binding fragment thereof includes a second antibody orfragment thereof that includes a second antibody or antigen-bindingfragment thereof (AB2) that binds a second, cancer-related target, wherethe AB2 is attached to a masking moiety (MM2) such that coupling of theMM2 reduces the ability of the AB2 to bind the second, cancer-relatedtarget.

In some embodiments, the T-cell engaging multispecific activatableantibody includes an OKT3 scFv or OKT3-derived scFv and a cancertargeting IgG antibody or antigen-binding fragment thereof, where atleast one of the OKT3 scFv or OKT3-derived scFv and/or the cancertargeting IgG antibody or antigen-binding portion thereof is masked. Insome embodiments, the OKT3 scFv or OKT3-derived scFv includes a firstantibody or antigen-binding fragment thereof (AB1) that binds CD3ε,where the AB1 is attached to a masking moiety (MM1) such that couplingof the MM1 reduces the ability of the AB1 to bind CD3ε. In someembodiments, the cancer targeting IgG antibody or antigen-bindingfragment thereof includes a second antibody or fragment thereof thatincludes a second antibody or antigen-binding fragment thereof (AB2)that binds a second, cancer-related target, where the AB2 is attached toa masking moiety (MM2) such that coupling of the MM2 reduces the abilityof the AB2 to bind the second, cancer-related target. In someembodiments, the OKT3 scFv or OKT3-derived scFv includes a firstantibody or antigen-binding fragment thereof (AB1) that binds CD3ε,where the AB1 is attached to a masking moiety (MM1) such that couplingof the MM1 reduces the ability of the AB1 to bind CD3ε, and the cancertargeting antibody IgG or antigen-binding fragment thereof includes asecond antibody or fragment thereof that includes a second antibody orantigen-binding fragment thereof (AB2) that binds a second,cancer-related target, where the AB2 is attached to a masking moiety(MM2) such that coupling of the MM2 reduces the ability of the AB2 tobind the second, cancer-related target.

In some embodiments, the T-cell engaging multispecific activatableantibody includes an anti-CD3 epsilon (CD3ε) scFv that is derived fromSP34, where at least one of the targeting antibody or antigen-bindingfragment thereof and/or the SP34 scFv or SP34-derived scFv is masked. Insome embodiments, the SP34 scFv or SP34-derived scFv includes a firstantibody or antigen-binding fragment thereof (AB1) that binds CD3ε,where the AB1 is attached to a masking moiety (MM1) such that couplingof the MM1 reduces the ability of the AB1 to bind CD3ε. In someembodiments, the targeting antibody or antigen-binding fragment thereofincludes a second antibody or fragment thereof that includes a secondantibody or antigen-binding fragment thereof (AB2) that binds a secondtarget, where the AB2 is attached to a masking moiety (MM2) such thatcoupling of the MM2 reduces the ability of the AB2 to bind the secondtarget. In some embodiments, the SP34 scFv or SP34-derived scFv includesa first antibody or antigen-binding fragment thereof (AB1) that bindsCD3ε, where the AB1 is attached to a masking moiety (MM1) such thatcoupling of the MM1 reduces the ability of the AB1 to bind CD3ε, and thetargeting antibody or antigen-binding fragment thereof includes a secondantibody or fragment thereof that includes a second antibody orantigen-binding fragment thereof (AB2) that binds a second target, wherethe AB2 is attached to a masking moiety (MM2) such that coupling of theMM2 reduces the ability of the AB2 to bind the second target.

In some embodiments, the T-cell engaging multispecific activatableantibody includes an SP34 scFv or SP34-derived scFv and a cancertargeting antibody or antigen-binding fragment thereof, where at leastone of the SP34 scFv or SP34-derived scFv and/or the cancer targetingantibody or antigen-binding portion thereof is masked. In someembodiments, the SP34 scFv or SP34-derived scFv includes a firstantibody or antigen-binding fragment thereof (AB1) that binds CD3ε,where the AB1 is attached to a masking moiety (MM1) such that couplingof the MM1 reduces the ability of the AB1 to bind CD3ε. In someembodiments, the cancer targeting antibody or antigen-binding fragmentthereof includes a second antibody or fragment thereof that includes asecond antibody or antigen-binding fragment thereof (AB2) that binds asecond, cancer-related target, where the AB2 is attached to a maskingmoiety (MM2) such that coupling of the MM2 reduces the ability of theAB2 to bind the second, cancer-related target. In some embodiments, theSP34 scFv or SP34-derived scFv includes a first antibody orantigen-binding fragment thereof (AB1) that binds CD3ε, where the AB1 isattached to a masking moiety (MM1) such that coupling of the MM1 reducesthe ability of the AB1 to bind CD3ε, and the cancer targeting antibodyor antigen-binding fragment thereof includes a second antibody orfragment thereof that includes a second antibody or antigen-bindingfragment thereof (AB2) that binds a second, cancer-related target, wherethe AB2 is attached to a masking moiety (MM2) such that coupling of theMM2 reduces the ability of the AB2 to bind the second, cancer-relatedtarget.

In some embodiments, the T-cell engaging multispecific activatableantibody includes an SP34 scFv or SP34-derived scFv and a cancertargeting IgG antibody or antigen-binding fragment thereof, where atleast one of the SP34 scFv or SP34-derived scFv and/or the cancertargeting IgG antibody or antigen-binding portion thereof is masked. Insome embodiments, the SP34 scFv or SP34-derived scFv includes a firstantibody or antigen-binding fragment thereof (AB1) that binds CD3ε,where the AB1 is attached to a masking moiety (MM1) such that couplingof the MM1 reduces the ability of the AB1 to bind CD3ε. In someembodiments, the cancer targeting IgG antibody or antigen-bindingfragment thereof includes a second antibody or fragment thereof thatincludes a second antibody or antigen-binding fragment thereof (AB2)that binds a second, cancer-related target, where the AB2 is attached toa masking moiety (MM2) such that coupling of the MM2 reduces the abilityof the AB2 to bind the second, cancer-related target. In someembodiments, the SP34 scFv or SP34-derived scFv includes a firstantibody or antigen-binding fragment thereof (AB1) that binds CD3ε,where the AB1 is attached to a masking moiety (MM1) such that couplingof the MM1 reduces the ability of the AB1 to bind CD3ε, and the cancertargeting antibody IgG or antigen-binding fragment thereof includes asecond antibody or fragment thereof that includes a second antibody orantigen-binding fragment thereof (AB2) that binds a second,cancer-related target, where the AB2 is attached to a masking moiety(MM2) such that coupling of the MM2 reduces the ability of the AB2 tobind the second, cancer-related target.

In some embodiments, the T-cell engaging multispecific activatableantibody includes an anti-CD3 epsilon (CD3ε) scFv that includes thefollowing complementarity determining region (CD3ε) sequences: a VH CDR1sequence that includes at least the amino acid sequence TYAMN (SEQ IDNO: 53); a VH CD2 sequence that includes at least the amino acidsequence RIRSKYNNYATYYADSVKD (SEQ ID NO: 54); a VH CDR3 sequence thatincludes at least the amino acid sequence HGNFGNSYVSWFAY (SEQ ID NO:55), a VL CDR1 sequence that includes at least the amino acid sequenceRSSTGAVTTSNYAN (SEQ ID NO: 56); a VL CDR2 sequence that includes atleast the amino acid sequence GTNKRAP (SEQ ID NO: 57); and a VL CDR3sequence that includes at least the amino acid sequence ALWYSNLWV (SEQID NO: 58), where at least one of the targeting antibody orantigen-binding fragment thereof and/or the anti-CD3ε scFv is masked. Insome embodiments, the anti-CD3ε scFv includes a first antibody orantigen-binding fragment thereof (AB1) having the CD3ε sequences set forthe above, i.e., SEQ ID NOs: 53, 54, 55, 56, 57, and 58, where the AB1is attached to a masking moiety (MM1) such that coupling of the MM1reduces the ability of the AB1 to bind CD3ε. In some embodiments, thetargeting antibody or antigen-binding fragment thereof includes a secondantibody or fragment thereof that includes a second antibody orantigen-binding fragment thereof (AB2) that binds a second target, wherethe AB2 is attached to a masking moiety (MM2) such that coupling of theMM2 reduces the ability of the AB2 to bind the second target. In someembodiments, the anti-CD3ε scFv includes a first antibody orantigen-binding fragment thereof (AB1) that includes the CD3ε sequencesof SEQ ID NOs: 53, 54, 55, 56, 57, and 58, where the AB1 is attached toa masking moiety (MM1) such that coupling of the MM1 reduces the abilityof the AB1 to bind CD3ε, and the targeting antibody or antigen-bindingfragment thereof includes a second antibody or fragment thereof thatincludes a second antibody or antigen-binding fragment thereof (AB2)that binds a second target, where the AB2 is attached to a maskingmoiety (MM2) such that coupling of the MM2 reduces the ability of theAB2 to bind the second target.

In some embodiments, the T-cell engaging multispecific activatableantibody includes an anti-CD3ε scFv that includes the CD3ε sequences ofSEQ ID NOs: 53, 54, 55, 56, 57, and 58 and a cancer targeting antibodyor antigen-binding fragment thereof, where at least one of the anti-CD3εscFv and/or the cancer targeting antibody or antigen-binding portionthereof is masked. In some embodiments, the anti-CD3ε scFv includes afirst antibody or antigen-binding fragment thereof (AB1) that includesthe CD3ε sequences of SEQ ID NOs: 53, 54, 55, 56, 57, and 58, where theAB1 is attached to a masking moiety (MM1) such that coupling of the MM1reduces the ability of the AB1 to bind CD3ε. In some embodiments, thecancer targeting antibody or antigen-binding fragment thereof includes asecond antibody or fragment thereof that includes a second antibody orantigen-binding fragment thereof (AB2) that binds a second,cancer-related target, where the AB2 is attached to a masking moiety(MM2) such that coupling of the MM2 reduces the ability of the AB2 tobind the second, cancer-related target. In some embodiments, theanti-CD3ε scFv includes a first antibody or antigen-binding fragmentthereof (AB1) that includes the CD3ε sequences of SEQ ID NOs: 53, 54,55, 56, 57, and 58, where the AB1 is attached to a masking moiety (MM1)such that coupling of the MM1 reduces the ability of the AB1 to bindCD3ε, and the cancer targeting antibody or antigen-binding fragmentthereof includes a second antibody or fragment thereof that includes asecond antibody or antigen-binding fragment thereof (AB2) that binds asecond, cancer-related target, where the AB2 is attached to a maskingmoiety (MM2) such that coupling of the MM2 reduces the ability of theAB2 to bind the second, cancer-related target.

In some embodiments, the T-cell engaging multispecific activatableantibody includes an anti-CD3εd scFv that includes the CD3ε sequences ofSEQ ID NOs: 53, 54, 55, 56, 57, and 58 and a cancer targeting IgGantibody or antigen-binding fragment thereof, where at least one of theanti-CD3ε scFv and/or the cancer targeting IgG antibody orantigen-binding portion thereof is masked. In some embodiments, theanti-CD3ε scFv includes a first antibody or antigen-binding fragmentthereof (AB1) that includes the CD3ε sequences of SEQ ID NOs: 53, 54,55, 56, 57, and 58, where the AB1 is attached to a masking moiety (MM1)such that coupling of the MM1 reduces the ability of the AB1 to bindCD3ε. In some embodiments, the cancer targeting IgG antibody orantigen-binding fragment thereof includes a second antibody or fragmentthereof that includes a second antibody or antigen-binding fragmentthereof (AB2) that binds a second, cancer-related target, where the AB2is attached to a masking moiety (MM2) such that coupling of the MM2reduces the ability of the AB2 to bind the second, cancer-relatedtarget. In some embodiments, the anti-CD3ε scFv includes a firstantibody or antigen-binding fragment thereof (AB1) that includes theCD3ε sequences of SEQ ID NOs: 53, 54, 55, 56, 57, and 58, where the AB1is attached to a masking moiety (MM1) such that coupling of the MM1reduces the ability of the AB1 to bind CD3ε, and the cancer targetingantibody IgG or antigen-binding fragment thereof includes a secondantibody or fragment thereof that includes a second antibody orantigen-binding fragment thereof (AB2) that binds a second,cancer-related target, where the AB2 is attached to a masking moiety(MM2) such that coupling of the MM2 reduces the ability of the AB2 tobind the second, cancer-related target.

In some embodiments, the multi-antigen targeting antibodies and/ormulti-antigen targeting activatable antibodies include at least a firstantibody or antigen-binding fragment thereof that binds a first targetand/or first epitope and a second antibody or antigen-binding fragmentthereof that binds a second target and/or a second epitope, where atleast one of the targets and/or epitopes is CD3ε. In some embodiments,the multi-antigen targeting antibodies and/or multi-antigen targetingactivatable antibodies bind two or more different targets, where atleast one target is CD3ε. In some embodiments, the multi-antigentargeting antibodies and/or multi-antigen targeting activatableantibodies bind two or more different epitopes on CD3ε. In someembodiments, the multi-antigen targeting antibodies and/or multi-antigentargeting activatable antibodies bind a combination of two or moredifferent targets and two or more different epitopes on the same target,where at least one of the targets and/or epitopes is CD3ε.

Various embodiments of multispecific activatable antibodies of thedisclosure are shown in FIGS. 5A-5D, 6A-6F, 7A-7J, 8A-8J, 9A-9J,10A-10J, 11A-11J, and 12A-12D. In some embodiments, a multispecificactivatable antibody comprising an IgG has the IgG variable domainsmasked. In some embodiments, a multispecific activatable antibodycomprising a scFv has the scFv domains masked. In some embodiments, amultispecific activatable antibody has both IgG variable domains andscFv domains, where at least one of the IgG variable domains is coupledto a masking moiety. In some embodiments, a multispecific activatableantibody has both IgG variable domains and scFv domains, where at leastone of the scFv domains is coupled to a masking moiety. In someembodiments, a multispecific activatable antibody has both IgG variabledomains and scFv domains, where at least one of the IgG variable domainsis coupled to a masking moiety and at least one of the scFv domains iscoupled to a masking moiety. In some embodiments, a multispecificactivatable antibody has both IgG variable domains and scFv domains,where each of the IgG variable domains and the scFv domains is coupledto its own masking moiety. In some embodiments, one antibody domain of amultispecific activatable antibody has specificity for a target antigenand another antibody domain has specificity for a T-cell surfaceantigen. In some embodiments, one antibody domain of a multispecificactivatable antibody has specificity for a target antigen and anotherantibody domain has specificity for another target antigen. In someembodiments, one antibody domain of a multispecific activatable antibodyhas specificity for an epitope of a target antigen and another antibodydomain has specificity for another epitope of the target antigen.

In a multispecific activatable antibody, a scFv can be fused to thecarboxyl terminus of the heavy chain of an IgG activatable antibody, tothe carboxyl terminus of the light chain of an IgG activatable antibody,or to the carboxyl termini of both the heavy and light chains of an IgGactivatable antibody. In a multispecific activatable antibody, a scFvcan be fused to the amino terminus of the heavy chain of an IgGactivatable antibody, to the amino terminus of the light chain of an IgGactivatable antibody, or to the amino termini of both the heavy andlight chains of an IgG activatable antibody. In a multispecificactivatable antibody, a scFv can be fused to any combination of one ormore carboxyl termini and one or more amino termini of an IgGactivatable antibody. In some embodiments, a masking moiety (MM) linkedto a cleavable moiety (CM) is attached to and masks an antigen bindingdomain of the IgG. In some embodiments, a masking moiety (MM) linked toa cleavable moiety (CM) is attached to and masks an antigen bindingdomain of at least one scFv. In some embodiments, a masking moiety (MM)linked to a cleavable moiety (CM) is attached to and masks an antigenbinding domain of an IgG and a masking moiety (MM) linked to a cleavablemoiety (CM) is attached to and masks an antigen binding domain of atleast one scFv.

In some embodiments, a single chain variable domain, specific forbinding CD3ε is fused to the carboxyl terminus of a fully human IgG1antibody (targeting antibody) that binds to a cell surface antigen.Fusion of the scFv can be to the carboxyl terminus of the heavy chain,to the carboxyl terminus of the light chain or to both chains. In someembodiments, a single chain variable domain, specific for binding CD3εis fused to the amino terminus of a fully human IgG1 antibody (targetingantibody) that binds to a cell surface antigen. Fusion of the scFv canbe to the amino terminus of the heavy chain, to the amino terminus ofthe light chain or to both chains. The fusions are constructed as asingle genetic construct and expressed in cells in culture. Thetargeting antibody can be specific for binding to one or more tumorsurface antigens, or any cell targeted for depletion. The scFv can bespecific for the same or different antigens.

Other examples of multispecific activatable antibody structures include,but are not limited to, the following:(VL-CL)₂:(VH-CH1-CH2-CH3-L4-VH*-L3-VL*-L2-CM-L1-MM)₂;(VL-CL)₂:(VH-CH1-CH2-CH3-L4-VL*-L3-VH*-L2-CM-L1-MM)₂;(MM-L1-CM-L2-VL-CL)₂:(VH-CH1-CH2-CH3-L4-VH*-L3-VL*)₂;(MM-L1-CM-L2-VL-CL)₂:(VH-CH1-CH2-CH3-L4-VL*-L3-VH*)₂;(VL-CL)₂:(MM-L1-CM-L2-VL*-L3-VH*-L4-VH-CH1-CH2-CH3)₂;(VL-CL)₂:(MM-L1-CM-L2-VH*-L3-VL*-L4-VH-CH1-CH2-CH3)₂;(MM-L1-CM-L2-VL-CL)₂:(VL*-L3-VH*-L4-VH-CH1-CH2-CH3)₂;(MM-L1-CM-L2-VL-CL)₂:(VH*-L3-VL*-L4-VH-CH1-CH2-CH3)₂;(VL-CL-L4-VH*-L3-VL*-L2-CM-L1-MM)₂:(VH-CH1-CH2-CH3)₂;(VL-CL-L4-VL*-L3-VH*-L2-CM-L1-MM)₂:(VH-CH1-CH2-CH3)₂;(MM-L1-CM-L2-VL*-L3-VH*-L4-VL-CL)₂:(VH-CH1-CH2-CH3)₂;(MM-L1-CM-L2-VH*-L3-VL*-L4-VL-CL)₂:(VH-CH1-CH2-CH3)₂;(VL-CL-L4-VH*-L3-VL*-L2-CM-L1-MM)₂:(MM-L1-CM-L2-VL*-L3-VH*-L4-VH-CH1-CH2-CH3)₂;(VL-CL-L4-VH*-L3-VL*-L2-CM-L1-MM)₂:(MM-L1-CM-L2-VH*-L3-VL*-L4-VH-CH1-CH2-CH3)₂;(VL-CL-L4-VL*-L3-VH*-L2-CM-L1-MM)₂:(MM-L1-CM-L2-VL*-L3-VH*-L4-VH-CH1-CH2-CH3)₂;(VL-CL-L4-VL*-L3-VH*-L2-CM-L1-MM)₂:(MM-L1-CM-L2-VH*-L3-VL*-L4-VH-CH1-CH2-CH3)₂; (VL-CL-L4-VH*-L3-VL*)₂:(MM-L1-CM-L2-VL*-L3-VH*-L4-VH-CH1-CH2-CH3)₂; (VL-CL-L4-VH*-L3-VL*)₂:(MM-L1-CM-L2-VH*-L3-VL*-L4-VH-CH1-CH2-CH3)₂; (VL-CL-L4-VL*-L3-VH*)₂:(MM-L1-CM-L2-VL*-L3-VH*-L4-VH-CH1-CH2-CH3)₂; (VL-CL-L4-VL*-L3-VH*)₂:(MM-L1-CM-L2-VH*-L3-VL*-L4-VH-CH1-CH2-CH3)2;(VL-CL-L4-VH*-L3-VL*-L2-CM-L1-MM)₂: (VL*-L3-VH*-L4-VH-CH1-CH2-CH3)₂;(VL-CL-L4-VH*-L3-VL*-L2-CM-L1-MM)₂: (VH*-L3-VL*-L4-VH-CH1-CH2-CH3)₂;(VL-CL-L4-VL*-L3-VH*-L2-CM-L1-MM)₂: (VL*-L3-VH*-L4-VH-CH1-CH2-CH3)₂; or(VL-CL-L4-VL*-L3-VH*-L2-CM-L1-MM)₂: (VH*-L3-VL*-L4-VH-CH1-CH2-CH3)₂,wherein: VL and VH represent the light and heavy variable domains of thefirst specificity, contained in the IgG; VL* and VH* represent thevariable domains of the second specificity, contained in the scFv; L1 isa linker peptide connecting the masking moiety (MM) and the cleavablemoiety (CM); L2 is a linker peptide connecting the cleavable moiety(CM), and the antibody; L3 is a linker peptide connecting the variabledomains of the scFv; L4 is a linker peptide connecting the antibody ofthe first specificity to the antibody of the second specificity; CL isthe light-chain constant domain; and CH1, CH2, CH3 are the heavy chainconstant domains. The first and second specificities may be toward anyantigen or epitope.

In some embodiments of a T-cell engaging multispecific activatableantibody, one antigen is typically CD3ε and the other is an antigenpresent on the surface of a tumor cell or other cell type associatedwith disease, such as, but not limited to, any target listed in Table 1,such as, but not limited to, EGFR, erbB2, EpCAM, Jagged, PD-L1, B7H3, orCD71 (transferrin receptor).

In some embodiments, the targeting antibody includes an anti-EGFRantibody. In some embodiments, the targeting antibody includes C225v5,which is specific for binding to EGFR. In some embodiments, thetargeting antibody includes C225, which is specific for binding to EGFR.In some embodiments, the targeting antibody includes C225v4, which isspecific for binding to EGFR. In some embodiments, the targetingantibody includes C225v6, which is specific for binding to EGFR. In someembodiments, the targeting antibody includes an anti-Jagged antibody. Insome embodiments, the targeting antibody includes 4D11, which isspecific for binding to human and mouse Jagged 1 and Jagged 2. In someembodiments, the targeting antibody includes 4D11v2, which is specificfor binding to human and mouse Jagged 1 and Jagged 2.

In some embodiments, the targeting antibody can be in the form anactivatable antibody. In some embodiments, the scFv(s) can be in theform of a Pro-scFv (see, e.g., WO 2009/025846, WO 2010/081173).

In some embodiments, the multispecific antibodies and/or multispecificactivatable antibodies provided herein include at least a first antibodyor antigen binding fragment thereof (AB1) that specifically binds CD3εand that contains a combination of a VH CDR1 sequence, a VH CDR2sequence, and a VH CDR3 sequence, wherein at least one of the VH CDR1sequence, the VH CDR2 sequence, and the VH CDR3 sequence is selectedfrom a VH CDR1 sequence that includes at least the amino acid sequenceTYAMN (SEQ ID NO: 53); a VH CD2 sequence that includes at least theamino acid sequence RIRSKYNNYATYYADSVKD (SEQ ID NO: 54); a VH CDR3sequence that includes at least the amino acid sequence HGNFGNSYVSWFAY(SEQ ID NO: 55), and combinations thereof.

In some embodiments, the multispecific antibodies and/or multispecificactivatable antibodies provided herein include at least a first antibodyor antigen binding fragment thereof (AB1) that specifically binds CD3εand that contains a combination of a VL CDR1 sequence, a VL CDR2sequence, and a VL CDR3 sequence, wherein at least one of the VL CDR1sequence, the VL CDR2 sequence, and the VL CDR3 sequence is selectedfrom a VL CDR1 sequence that includes at least the amino acid sequenceRSSTGAVTTSNYAN (SEQ ID NO: 56); a VL CDR2 sequence that includes atleast the amino acid sequence GTNKRAP (SEQ ID NO: 57); and a VL CDR3sequence that includes at least the amino acid sequence ALWYSNLWV (SEQID NO: 58), and combinations thereof.

In some embodiments, the multispecific antibodies and/or multispecificactivatable antibodies provided herein include at least a first antibodyor antigen binding fragment thereof (AB1) that specifically binds CD3εand that contains a VH CDR1 sequence that includes at least the aminoacid sequence TYAMN (SEQ ID NO: 53); a VH CD2 sequence that includes atleast the amino acid sequence RIRSKYNNYATYYADSVKD (SEQ ID NO: 54); a VHCDR3 sequence that includes at least the amino acid sequenceHGNFGNSYVSWFAY (SEQ ID NO: 55), a VL CDR1 sequence that includes atleast the amino acid sequence RSSTGAVTTSNYAN (SEQ ID NO: 56); a VL CDR2sequence that includes at least the amino acid sequence GTNKRAP (SEQ IDNO: 57); and a VL CDR3 sequence that includes at least the amino acidsequence ALWYSNLWV (SEQ ID NO: 58).

In some embodiments, the multispecific antibodies and/or multispecificactivatable antibodies provided herein include at least a first antibodyor antigen binding fragment thereof (AB1) that specifically binds CD3εand that contains a VH CDR1 sequence that is at least 90%, 91%, 92%,93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical to the amino acidsequence TYAMN (SEQ ID NO: 53); a VH CD2 sequence that is at least 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical to theamino acid sequence RIRSKYNNYATYYADSVKD (SEQ ID NO: 54); a VH CDR3sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%,99% or more identical to the amino acid sequence HGNFGNSYVSWFAY (SEQ IDNO: 55), a VL CDR1 sequence that is at least 90%, 91%, 92%, 93%, 94%,95%, 96%, 97%, 98%, 99% or more identical to the amino acid sequenceRSSTGAVTTSNYAN (SEQ ID NO: 56); a VL CDR2 sequence that is at least 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical to theamino acid sequence GTNKRAP (SEQ ID NO: 57); and a VL CDR3 sequence thatis at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or moreidentical to the amino acid sequence ALWYSNLWV (SEQ ID NO: 58).

In some embodiments, the multispecific antibodies and/or multispecificactivatable antibodies provided herein include at least a first antibodyor antigen binding fragment thereof (AB1) that specifically binds aJagged target, e.g., Jagged 1 and/or Jagged 2, and that contains acombination of a VH CDR1 sequence, a VH CDR2 sequence, and a VH CDR3sequence, wherein at least one of the VH CDR1 sequence, the VH CDR2sequence, and the VH CDR3 sequence is selected from a VH CDR1 sequencethat includes at least the amino acid sequence SYAMS (SEQ ID NO: 88); aVH CD2 sequence that includes at least the amino acid sequenceSIDPEGRQTYYADSVKG (SEQ ID NO: 89); and a VH CDR3 sequence that includesat least the amino acid sequence DIGGRSAFDY (SEQ ID NO: 90), andcombinations thereof.

In some embodiments, the multispecific antibodies and/or multispecificactivatable antibodies provided herein include at least a first antibodyor antigen binding fragment thereof (AB1) that specifically binds aJagged target, e.g., Jagged 1 and/or Jagged 2, and that contains acombination of a VL CDR1 sequence, a VL CDR2 sequence, and a VL CDR3sequence, wherein at least one of the VL CDR1 sequence, the VL CDR2sequence, and the VL CDR3 sequence is selected from a VL CDR1 sequencethat includes at least the amino acid sequence RASQSISSY (SEQ ID NO:91); a VL CDR2 sequence that includes at least the amino acid sequenceAASSLQS (SEQ ID NO: 92); and a VL CDR3 sequence that includes at leastthe amino acid sequence QQTVVAPPL (SEQ ID NO: 93), and combinationsthereof.

In some embodiments, the multispecific antibodies and/or multispecificactivatable antibodies provided herein include at least a first antibodyor antigen binding fragment thereof (AB1) that specifically binds aJagged target, e.g., Jagged 1 and/or Jagged 2, and that contains acombination of a VH CDR1 sequence, a VH CDR2 sequence, and a VH CDR3sequence, wherein at least one of the VH CDR1 sequence, the VH CDR2sequence, and the VH CDR3 sequence is selected from a VH CDR1 sequencethat includes a sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%,96%, 97%, 98%, 99% or more identical to the amino acid sequence SYAMS(SEQ ID NO: 88); a VH CD2 sequence that includes a sequence that is atleast 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identicalto the amino acid sequence SIDPEGRQTYYADSVKG (SEQ ID NO: 89); and a VHCDR3 sequence that includes a sequence that is at least 90%, 91%, 92%,93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical to the amino acidsequence DIGGRSAFDY (SEQ ID NO: 90), and combinations thereof.

In some embodiments, the multispecific antibodies and/or multispecificactivatable antibodies provided herein include at least a first antibodyor antigen binding fragment thereof (AB1) that specifically binds aJagged target, e.g., Jagged 1 and/or Jagged 2, and that contains acombination of a VL CDR1 sequence, a VL CDR2 sequence, and a VL CDR3sequence, wherein at least one of the VL CDR1 sequence, the VL CDR2sequence, and the VL CDR3 sequence is selected from a VL CDR1 sequencethat includes a sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%,96%, 97%, 98%, 99% or more identical to the amino acid sequenceRASQSISSY (SEQ ID NO: 91); a VL CDR2 sequence that includes a sequencethat is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% ormore identical to the amino acid sequence AASSLQS (SEQ ID NO: 92); and aVL CDR3 sequence that includes a sequence that is at least 90%, 91%,92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical to the aminoacid sequence QQTVVAPPL (SEQ ID NO: 93), and combinations thereof.

In some embodiments, the multispecific antibodies and/or multispecificactivatable antibodies provided herein include at least a first antibodyor antigen binding fragment thereof (AB1) that specifically binds aJagged target, e.g., Jagged 1 and/or Jagged 2, and that contains acombination of a VH CDR1 sequence, a VH CDR2 sequence, a VH CDR3sequence, a VL CDR1 sequence, a VL CDR2 sequence, and a VL CDR3sequence, wherein the VH CDR1 sequence includes at least the amino acidsequence SYAMS (SEQ ID NO: 88); the VH CD2 sequence includes at leastthe amino acid sequence SIDPEGRQTYYADSVKG (SEQ ID NO: 89); the VH CDR3sequence includes at least the amino acid sequence DIGGRSAFDY (SEQ IDNO: 90); the VL CDR1 sequence includes at least the amino acid sequenceRASQSISSY (SEQ ID NO: 91); the VL CDR2 sequence includes at least theamino acid sequence AASSLQS (SEQ ID NO: 92); and the VL CDR3 sequenceincludes at least the amino acid sequence QQTVVAPPL (SEQ ID NO: 93).

In some embodiments, the multispecific antibodies and/or multispecificactivatable antibodies provided herein include at least a first antibodyor antigen binding fragment thereof (AB1) that specifically binds aJagged target, e.g., Jagged 1 and/or Jagged 2, and that contains acombination of a VH CDR1 sequence, a VH CDR2 sequence, a VH CDR3sequence, a VL CDR1 sequence, a VL CDR2 sequence, and a VL CDR3sequence, wherein the VH CDR1 sequence includes a sequence that is atleast 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identicalto the amino acid sequence SYAMS (SEQ ID NO: 88); the VH CD2 sequenceincludes a sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%,97%, 98%, 99% or more identical to the amino acid sequenceSIDPEGRQTYYADSVKG (SEQ ID NO: 89); the VH CDR3 sequence includes asequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%,99% or more identical to the amino acid sequence DIGGRSAFDY (SEQ ID NO:90); the VL CDR1 sequence includes a sequence that is at least 90%, 91%,92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical to the aminoacid sequence RASQSISSY (SEQ ID NO: 91); the VL CDR2 sequence includes asequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%,99% or more identical to the amino acid sequence AASSLQS (SEQ ID NO:92); and the VL CDR3 sequence includes a sequence that is at least 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical to theamino acid sequence QQTVVAPPL (SEQ ID NO: 93).

In some embodiments, the multispecific antibodies and/or multispecificactivatable antibodies provided herein include at least a first antibodyor antigen binding fragment thereof (AB1) that specifically bindsEpidermal Growth Factor Receptor (EGFR) and that contains a combinationof a VH CDR1 sequence, a VH CDR2 sequence, and a VH CDR3 sequence,wherein at least one of the VH CDR1 sequence, the VH CDR2 sequence, andthe VH CDR3 sequence is selected from a VH CDR1 sequence that includesat least the amino acid sequence NYGVH (SEQ ID NO: 94); a VH CD2sequence that includes at least the amino acid sequence VIWSGGNTDYNTPFTS(SEQ ID NO: 95); a VH CDR3 sequence that includes at least the aminoacid sequence ALTYYDYEFAY (SEQ ID NO: 96); and combinations thereof.

In some embodiments, the multispecific antibodies and/or multispecificactivatable antibodies provided herein include at least a first antibodyor antigen binding fragment thereof (AB1) that specifically binds EGFRand that contains a combination of a VL CDR1 sequence, a VL CDR2sequence, and a VL CDR3 sequence, wherein at least one of the VL CDR1sequence, the VL CDR2 sequence, and the VL CDR3 sequence is selectedfrom a VL CDR1 sequence that includes at least the amino acid sequenceRASQSIGTNIH (SEQ ID NO: 97); a VL CDR2 sequence that includes at leastthe amino acid sequence KYASESIS (SEQ ID NO: 98); and a VL CDR3 sequencethat includes at least the amino acid sequence QQNNNWPTT (SEQ ID NO:99), and combinations thereof.

In some embodiments, the multispecific antibodies and/or multispecificactivatable antibodies provided herein include at least a first antibodyor antigen binding fragment thereof (AB1) that specifically binds EGFRand that contains a combination of a VH CDR1 sequence, a VH CDR2sequence, and a VH CDR3 sequence, wherein at least one of the VH CDR1sequence, the VH CDR2 sequence, and the VH CDR3 sequence is selectedfrom a VH CDR1 sequence that includes a sequence that is at least 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical to theamino acid sequence NYGVH (SEQ ID NO: 94); a VH CD2 sequence thatincludes a sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%,97%, 98%, 99% or more identical to the amino acid sequenceVIWSGGNTDYNTPFTS (SEQ ID NO: 95); a VH CDR3 sequence that includes asequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%,99% or more identical to the amino acid sequence ALTYYDYEFAY (SEQ ID NO:96); and combinations thereof.

In some embodiments, the multispecific antibodies and/or multispecificactivatable antibodies provided herein include at least a first antibodyor antigen binding fragment thereof (AB1) that specifically binds EGFRand that contains a combination of a VL CDR1 sequence, a VL CDR2sequence, and a VL CDR3 sequence, wherein at least one of the VL CDR1sequence, the VL CDR2 sequence, and the VL CDR3 sequence is selectedfrom a VL CDR1 sequence that includes a sequence that is at least 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical to theamino acid sequence RASQSIGTNIH (SEQ ID NO: 97); a VL CDR2 sequence thatincludes a sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%,97%, 98%, 99% or more identical to the amino acid sequence KYASESIS (SEQID NO: 98); and a VL CDR3 sequence that includes a sequence that is atleast 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identicalto the amino acid sequence QQNNNWPTT (SEQ ID NO: 99), and combinationsthereof.

In some embodiments, the multispecific antibodies and/or multispecificactivatable antibodies provided herein include at least a first antibodyor antigen binding fragment thereof (AB1) that specifically binds EGFRand that contains a combination of a VH CDR1 sequence, a VH CDR2sequence, a VH CDR3 sequence, a VL CDR1 sequence, a VL CDR2 sequence,and a VL CDR3 sequence, wherein the VH CDR1 sequence includes at leastthe amino acid sequence NYGVH (SEQ ID NO: 94); the VH CD2 sequenceincludes at least the amino acid sequence VIWSGGNTDYNTPFTS (SEQ ID NO:95); the VH CDR3 sequence includes at least the amino acid sequenceALTYYDYEFAY (SEQ ID NO: 96); the VL CDR1 sequence includes at least theamino acid sequence RASQSIGTNIH (SEQ ID NO: 97); the VL CDR2 sequenceincludes at least the amino acid sequence KYASESIS (SEQ ID NO: 98); andthe VL CDR3 sequence includes at least the amino acid sequence QQNNNWPTT(SEQ ID NO: 99).

In some embodiments, the multispecific antibodies and/or multispecificactivatable antibodies provided herein include at least a first antibodyor antigen binding fragment thereof (AB1) that specifically binds EGFRand that contains a combination of a VH CDR1 sequence, a VH CDR2sequence, a VH CDR3 sequence, a VL CDR1 sequence, a VL CDR2 sequence,and a VL CDR3 sequence, wherein the VH CDR1 sequence includes a sequencethat is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% ormore identical to the amino acid sequence NYGVH (SEQ ID NO: 94); the VHCD2 sequence includes a sequence that is at least 90%, 91%, 92%, 93%,94%, 95%, 96%, 97%, 98%, 99% or more identical to the amino acidsequence VIWSGGNTDYNTPFTS (SEQ ID NO: 95); the VH CDR3 sequence includesa sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%,99% or more identical to the amino acid sequence ALTYYDYEFAY (SEQ ID NO:96); the VL CDR1 sequence includes a sequence that is at least 90%, 91%,92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical to the aminoacid sequence RASQSIGTNIH (SEQ ID NO: 97); the VL CDR2 sequence includesa sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%,99% or more identical to the amino acid sequence KYASESIS (SEQ ID NO:98); and the VL CDR3 sequence includes a sequence that is at least 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical to theamino acid sequence QQNNNWPTT (SEQ ID NO: 99).

In some embodiments, the multispecific antibodies and/or multispecificactivatable antibodies provided herein include at least a heavy chainamino acid sequence selected from the group consisting of thosesequences shown in the Examples provided herein. In some embodiments,the multispecific antibodies and/or multispecific activatable antibodiesprovided herein include at least a light chain amino acid sequenceselected from the group consisting of those sequences shown in theExamples provided herein. In some embodiments, the multispecificantibodies and/or multispecific activatable antibodies provided hereininclude at least a heavy chain amino acid sequence selected from thegroup consisting of those sequences shown in the Examples providedherein, and a light chain amino acid sequence selected from the groupconsisting of those sequences shown in the Examples provided herein.

In some embodiments, the multispecific antibodies and/or multispecificactivatable antibodies provided herein include at least a heavy chainamino acid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%,97%, 98%, 99% or more identical to an amino acid sequence selected fromthe group consisting of those sequences shown in the Examples providedherein. In some embodiments, the multispecific antibodies and/ormultispecific activatable antibodies provided herein include at least alight chain amino acid sequence that is at least 90%, 91%, 92%, 93%,94%, 95%, 96%, 97%, 98%, 99% or more identical to an amino acid sequenceselected from the group consisting of those sequences shown in theExamples provided herein. In some embodiments, the multispecificantibodies and/or multispecific activatable antibodies provided hereininclude at least a heavy chain amino acid sequence that is at least 90%,91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more identical to anamino acid sequence selected from the group consisting of thosesequences shown in the Examples provided herein, and a light chain aminoacid sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%,98%, 99% or more identical to an amino acid sequence selected from thegroup consisting of those sequences shown in the Examples providedherein.

In some embodiments, the multispecific antibody and/or multispecificactivatable antibody also includes an agent conjugated to the AB. Insome embodiments, the agent is a therapeutic agent. In some embodiments,the agent is a detectable moiety. In some embodiments, the detectablemoiety is a diagnostic agent. In some embodiments, the agent isconjugated to the multispecific via a linker. In some embodiments, thelinker is a cleavable linker. In some embodiments, the linker is anon-cleavable linker.

In some embodiments, the multispecific antibody and/or multispecificactivatable antibody also includes a detectable moiety. In someembodiments, the detectable moiety is a diagnostic agent.

In some embodiments, the multispecific antibody and/or multispecificactivatable antibody naturally contains one or more disulfide bonds. Insome embodiments, the multispecific antibody and/or multispecificactivatable antibody can be engineered to include one or more disulfidebonds.

The disclosure also provides an isolated nucleic acid molecule encodingat least a portion of a multispecific antibody and/or multispecificactivatable antibody described herein, such as for example, at least afirst nucleic acid encoding at least a portion of the heavy chain of themultispecific antibody and/or multispecific activatable antibody and asecond nucleic acid encoding at least a portion of the light chain ofthe multispecific antibody and/or multispecific activatable antibody, aswell as vectors that include these isolated nucleic acid sequences. Thedisclosure provides methods of producing a multispecific antibody byculturing a cell under conditions that lead to expression of theantibody, wherein the cell comprises such a nucleic acid molecule(s). Insome embodiments, the cell comprises such a vector.

The disclosure also provides multispecific activatable antibodies and/ormultispecific activatable antibody compositions that include at least afirst antibody or antigen-binding fragment thereof (AB1) thatspecifically binds a first target or first epitope and a second antibodyor antigen-biding fragment thereof (AB2) that binds a second target or asecond epitope, where at least AB1 is coupled or otherwise attached to amasking moiety (MM1), such that coupling of the MM1 reduces the abilityof AB1 to bind its target. In some embodiments, the MM1 is coupled toAB1 via a first cleavable moiety (CM1) sequence that includes asubstrate for a protease, for example, a protease that is produced by atumor that is in proximity to cells that express the target and/orproduced by a tumor that is co-localized with the target of AB1 at atreatment site or a diagnostic site in a subject. The multispecificactivatable antibodies provided herein are stable in circulation,activated at intended sites of therapy and/or diagnosis but not innormal, i.e., healthy tissue, and, when activated, exhibit binding tothe target of AB1 that is at least comparable to the corresponding,unmodified multispecific antibody.

In some embodiments, the multispecific activatable antibody comprises alinking peptide between the MM1 and the CM1.

In some embodiments, the multispecific activatable antibody comprises alinking peptide between the CM1 and the AB1.

In some embodiments, the activatable antibody comprises a first linkingpeptide (LP1) and a second linking peptide (LP2), and at least a portionof the multispecific activatable antibody in the uncleaved state has thestructural arrangement from N-terminus to C-terminus as follows:MM1-LP1-CM1-LP2-AB1 or AB1-LP2-CM1-LP1-MM1. In some embodiments, the twolinking peptides need not be identical to each other.

In some embodiments, at least one of LP1 or LP2 includes an amino acidsequence selected from the group consisting of (GS)_(n), (GGS)_(n),(GSGGS)_(n) (SEQ ID NO: 59) and (GGGS)_(n) (SEQ ID NO: 60), where n isan integer of at least one. In some embodiments, at least one of LP1 orLP2 includes an amino acid sequence selected from the group consistingof GGSG (SEQ ID NO: 61), GGSGG (SEQ ID NO: 62), GSGSG (SEQ ID NO: 63),GSGGG (SEQ ID NO: 64), GGGSG (SEQ ID NO: 65), and GSSSG (SEQ ID NO: 66).

In some embodiments, the multispecific activatable antibody includes atleast a first antibody or antigen-binding fragment thereof (AB1) thatspecifically binds a first target or first epitope and a second antibodyor antigen-binding fragment thereof (AB2) that specifically binds asecond target or second epitope. In some embodiments, each of the AB inthe multispecific activatable antibody is independently selected fromthe group consisting of a monoclonal antibody, domain antibody, singlechain, Fab fragment, a F(ab′)₂ fragment, a scFv, a scAb, a dAb, a singledomain heavy chain antibody, and a single domain light chain antibody.In some embodiments, each of the AB in the multispecific activatableantibody is a rodent (e.g., mouse or rat), chimeric, humanized or fullyhuman monoclonal antibody.

In some embodiments, each of the AB in the multispecific activatableantibody has a dissociation constant of about 100 nM or less for bindingto its corresponding target or epitope.

In some embodiments, the MM1 has a dissociation constant, i.e.,dissociation constant at an equilibrium state, K_(d) for binding to theAB that is greater than the K_(d) for binding of the AB to itscorresponding target or epitope.

In some embodiments, the MM1 has a K_(d) for binding to the AB that isno more than the K_(d) for binding of the AB to its corresponding targetor epitope.

In some embodiments, the MM1 has a K_(d) for binding to the AB that isno less than the K_(d) for binding of the AB to its corresponding targetor epitope.

In some embodiments, the MM1 has a K_(d) for binding to the AB that isapproximately equal to the K_(d) for binding of the AB to itscorresponding target or epitope.

In some embodiments, the MM1 has a K_(d) for binding to the AB that isless than the K_(d) for binding of the AB to its corresponding target orepitope.

In some embodiments, the MM1 has a K_(d) for binding to the AB that isno more than 2, 3, 4, 5, 10, 25, 50, 100, 250, 500, or 1,000 foldgreater than the K_(d) for binding of the AB to its corresponding targetor epitope. In some embodiments, the MM1 has a K_(d) for binding to theAB that is between 1-5, 2-5, 2-10, 5-10, 5-20, 5-50, 5-100, 10-100,10-1,000, 20-100, 20-1000, or 100-1,000 fold greater than the K_(d) forbinding of the AB to its corresponding target or epitope.

In some embodiments, the MM1 has an affinity for binding to the AB thatis less than the affinity of binding of the AB to its correspondingtarget or epitope.

In some embodiments, the MM1 has an affinity for binding to the AB thatis no more than the affinity of binding of the AB to its correspondingtarget or epitope.

In some embodiments, the MM1 has an affinity for binding to the AB thatis approximately equal of the affinity of binding of the AB to itscorresponding target or epitope.

In some embodiments, the MM1 has an affinity for binding to the AB thatis no less than the affinity of binding of the AB to its correspondingtarget or epitope.

In some embodiments, the MM1 has an affinity for binding to the AB thatis greater than the affinity of binding of the AB to its correspondingtarget or epitope.

In some embodiments, the MM1 has an affinity for binding to the AB thatis 2, 3, 4, 5, 10, 25, 50, 100, 250, 500, or 1,000 less than theaffinity of binding of the AB to its corresponding target or epitope. Insome embodiments, the MM1 has an affinity for binding to the AB that isbetween 1-5, 2-5, 2-10, 5-10, 5-20, 5-50, 5-100, 10-100, 10-1,000,20-100, 20-1000, or 100-1,000 fold less than the affinity of binding ofthe AB to its corresponding target or epitope. In some embodiments, theMM1 has an affinity for binding to the AB that is 2 to 20 fold less thanthe affinity of binding of the AB to its corresponding target orepitope. In some embodiments, a MM not covalently linked to the AB andat equimolar concentration to the AB does not inhibit the binding of theAB to its corresponding target or epitope.

In some embodiments, MM1 does not interfere or compete with itscorresponding AB for binding to the corresponding target or epitope whenthe multispecific activatable antibody is in a cleaved state.

In some embodiments, MM1 is a polypeptide of about 2 to 40 amino acidsin length. In some embodiments, each of the MM in the multispecificactivatable antibody is a polypeptide of no more than 40 amino acids inlength.

In some embodiments, MM1 has a polypeptide sequence that is differentfrom that of target of the corresponding AB.

In some embodiments, MM1 has a polypeptide sequence that is no more than50% identical to any natural binding partner of the corresponding AB. Insome embodiments, MM1 has a polypeptide sequence that is no more than25% identical to any natural binding partner of the corresponding AB. Insome embodiments, MM1 has a polypeptide sequence that is no more than10% identical to any natural binding partner of the corresponding AB.

In some embodiments, the coupling of MM1 reduces the ability of thecorresponding AB to bind its target or epitope such that thedissociation constant (K_(d)) of the AB when coupled to the MM1 towardsits corresponding target or epitope is at least 20 times greater thanthe K_(d) of the AB when not coupled to the MM1 towards itscorresponding target or epitope.

In some embodiments, the coupling of MM1 reduces the ability of thecorresponding AB to bind its target or epitope such that thedissociation constant (K_(d)) of the AB when coupled to the MM1 towardsits corresponding target or epitope is at least 40 times greater thanthe K_(d) of the AB when not coupled to the MM1 towards itscorresponding target or epitope.

In some embodiments, the coupling of MM1 reduces the ability of thecorresponding AB to bind its target or epitope such that thedissociation constant (K_(d)) of the AB when coupled to the MM1 towardsits corresponding target or epitope is at least 100 times greater thanthe K_(d) of the AB when not coupled to the MM1 towards itscorresponding target or epitope.

In some embodiments, the coupling of MM1 reduces the ability of thecorresponding AB to bind its target or epitope such that thedissociation constant (K_(d)) of the AB when coupled to the MM1 towardsits corresponding target or epitope is at least 1000 times greater thanthe K_(d) of the AB when not coupled to the MM1 towards itscorresponding target or epitope.

In some embodiments, the coupling of MM1 reduces the ability of thecorresponding AB to bind its target or epitope such that thedissociation constant (K_(d)) of the AB when coupled to the MM1 towardsits corresponding target or epitope is at least 10,000 times greaterthan the K_(d) of the AB when not coupled to the MM1 towards itscorresponding target or epitope.

In some embodiments, MM1 is an amino acid sequence selected from a MMshown in the Examples provided herein.

In some embodiments, the multispecific activatable antibody includes atleast a second masking moiety (MM2) that inhibits the binding of the AB2to its target when the multispecific activatable antibody is in anuncleaved state, and a second cleavable moiety (CM2) coupled to the AB2,wherein the CM2 is a polypeptide that functions as a substrate for asecond protease. In some embodiments, CM2 is a polypeptide of no morethan 15 amino acids long. In some embodiments, the second protease isproduced by a tumor that is in proximity to cells that express thetarget and/or produced by a tumor that is co-localized with the secondtarget or epitope in a tissue, and wherein the second protease cleavesthe CM2 in the multispecific activatable antibody when the multispecificactivatable antibody is exposed to the second protease. In someembodiments, the first protease and the second protease are produced bya tumor that is in proximity to cells that express the target and/orproduced by a tumor that is co-localized with the first target orepitope and the second target or epitope in a tissue. In someembodiments, the first protease and the second protease are the sameprotease. In some embodiments, CM1 and CM2 are different substrates forthe same protease. In some embodiments, the protease is selected fromthe group consisting of those shown in Table 3. In some embodiments, thefirst protease and the second protease are different proteases. In someembodiments, the first protease and the second protease are differentproteases selected from the group consisting of those shown in Table 3.

In some embodiments, each of the MM in the multispecific activatableantibody has a dissociation constant, i.e., dissociation constant at anequilibrium state, K_(d) for binding to the AB that is greater than theK_(d) for binding of the AB to its corresponding target or epitope.

In some embodiments, each of the MM in the multispecific activatableantibody has a K_(d) for binding to the AB that is no more than theK_(d) for binding of the AB to its corresponding target or epitope.

In some embodiments, each of the MM in the multispecific activatableantibody has a K_(d) for binding to the AB that is no less than theK_(d) for binding of the AB to its corresponding target or epitope.

In some embodiments, each of the MM in the multispecific activatableantibody has a K_(d) for binding to the AB that is approximately equalto the K_(d) for binding of the AB to its corresponding target orepitope.

In some embodiments, each of the MM in the multispecific activatableantibody has a K_(d) for binding to the AB that is less than the K_(d)for binding of the AB to its corresponding target or epitope.

In some embodiments, each of the MM in the multispecific activatableantibody has a IQ for binding to the AB that is no more than 2, 3, 4, 5,10, 25, 50, 100, 250, 500, or 1,000 fold greater than the K_(d) forbinding of the AB to its corresponding target or epitope. In someembodiments, each of the MM in the multispecific activatable antibodyhas a K_(d) for binding to the AB that is between 1-5, 2-5, 2-10, 5-10,5-20, 5-50, 5-100, 10-100, 10-1,000, 20-100, 20-1000, or 100-1,000 foldgreater than the K_(d) for binding of the AB to its corresponding targetor epitope.

In some embodiments, each of the MM in the multispecific activatableantibody has an affinity for binding to the AB that is less than theaffinity of binding of the AB to its corresponding target or epitope.

In some embodiments, each of the MM in the multispecific activatableantibody has an affinity for binding to the AB that is no more than theaffinity of binding of the AB to its corresponding target or epitope.

In some embodiments, each of the MM in the multispecific activatableantibody has an affinity for binding to the AB that is approximatelyequal of the affinity of binding of the AB to its corresponding targetor epitope.

In some embodiments, each of the MM in the multispecific activatableantibody has an affinity for binding to the AB that is no less than theaffinity of binding of the AB to its corresponding target or epitope.

In some embodiments, each of the MM in the multispecific activatableantibody has an affinity for binding to the AB that is greater than theaffinity of binding of the AB to its corresponding target or epitope.

In some embodiments, each of the MM in the multispecific activatableantibody has an affinity for binding to the AB that is 2, 3, 4, 5, 10,25, 50, 100, 250, 500, or 1,000 less than the affinity of binding of theAB to its corresponding target or epitope. In some embodiments, each ofthe MM in the multispecific activatable antibody has an affinity forbinding to the AB that is between 1-5, 2-5, 2-10, 5-10, 5-20, 5-50,5-100, 10-100, 10-1,000, 20-100, 20-1000, or 100-1,000 fold less thanthe affinity of binding of the AB to its corresponding target orepitope. In some embodiments, each of the MM in the multispecificactivatable antibody has an affinity for binding to the AB that is 2 to20 fold less than the affinity of binding of the AB to its correspondingtarget or epitope. In some embodiments, a MM not covalently linked tothe AB and at equimolar concentration to the AB does not inhibit thebinding of the AB to its corresponding target or epitope.

In some embodiments, each of the MM in the multispecific activatableantibody does not interfere or compete with its corresponding AB forbinding to the corresponding target or epitope when the multispecificactivatable antibody is in a cleaved state.

In some embodiments, each of the MM in the multispecific activatableantibody is a polypeptide of about 2 to 40 amino acids in length. Insome embodiments, each of the MM in the multispecific activatableantibody is a polypeptide of no more than 40 amino acids in length.

In some embodiments, each of the MM in the multispecific activatableantibody has a polypeptide sequence that is different from that oftarget of the corresponding AB.

In some embodiments, each of the MM in the multispecific activatableantibody has a polypeptide sequence that is no more than 50% identicalto any natural binding partner of the corresponding AB. In someembodiments, each of the MM in the multispecific activatable antibodyhas a polypeptide sequence that is no more than 25% identical to anynatural binding partner of the corresponding AB. In some embodiments,each of the MM in the multispecific activatable antibody has apolypeptide sequence that is no more than 10% identical to any naturalbinding partner of the corresponding AB.

In some embodiments, the coupling of each of the MM reduces the abilityof the corresponding AB to bind its target or epitope such that thedissociation constant (K_(d)) of the AB when coupled to the MM towardsits corresponding target or epitope is at least 20 times greater thanthe K_(d) of the AB when not coupled to the MM towards its correspondingtarget or epitope.

In some embodiments, the coupling of each of the MM reduces the abilityof the corresponding AB to bind its target or epitope such that thedissociation constant (K_(d)) of the AB when coupled to the MM towardsits corresponding target or epitope is at least 40 times greater thanthe K_(d) of the AB when not coupled to the MM towards its correspondingtarget or epitope.

In some embodiments, the coupling of each of the MM reduces the abilityof the corresponding AB to bind its target or epitope such that thedissociation constant (K_(d)) of the AB when coupled to the MM towardsits corresponding target or epitope is at least 100 times greater thanthe K_(d) of the AB when not coupled to the MM towards its correspondingtarget or epitope.

In some embodiments, the coupling of each of the MM reduces the abilityof the corresponding AB to bind its target or epitope such that thedissociation constant (K_(d)) of the AB when coupled to the MM towardsits corresponding target or epitope is at least 1000 times greater thanthe K_(d) of the AB when not coupled to the MM towards its correspondingtarget or epitope.

In some embodiments, the coupling of each of the MM reduces the abilityof the corresponding AB to bind its target or epitope such that thedissociation constant (K_(d)) of the AB when coupled to the MM towardsits corresponding target or epitope is at least 10,000 times greaterthan the K_(d) of the AB when not coupled to the MM towards itscorresponding target or epitope.

In some embodiments, the coupling of each of the MM reduces the abilityof the corresponding AB to bind its target or epitope such that thedissociation constant (K_(d)) of the AB1 when coupled to the MM1 is atleast 20 times greater than the K_(d) of the AB2 when coupled to theMM2. In some embodiments, the coupling of each of the MM reduces theability of the corresponding AB to bind its target or epitope such thatthe dissociation constant (K_(d)) of the AB1 when coupled to the MM1 isat least 20 times lower than the K_(d) of the AB2 when coupled to theMM2.

In some embodiments, the coupling of each of the MM reduces the abilityof the corresponding AB to bind its target or epitope such that thedissociation constant (K_(d)) of the AB1 when coupled to the MM1 is atleast 40 times greater than the K_(d) of the AB2 when coupled to theMM2. In some embodiments, the coupling of each of the MM reduces theability of the corresponding AB to bind its target or epitope such thatthe dissociation constant (K_(d)) of the AB1 when coupled to the MM1 isat least 40 times lower than the K_(d) of the AB2 when coupled to theMM2.

In some embodiments, the coupling of each of the MM reduces the abilityof the corresponding AB to bind its target or epitope such that thedissociation constant (K_(d)) of the AB1 when coupled to the MM1 is atleast 100 times greater than the K_(d) of the AB2 when coupled to theMM2. In some embodiments, the coupling of each of the MM reduces theability of the corresponding AB to bind its target or epitope such thatthe dissociation constant (K_(d)) of the AB1 when coupled to the MM1 isat least 100 times lower than the K_(d) of the AB2 when coupled to theMM2.

In some embodiments, the coupling of each of the MM reduces the abilityof the corresponding AB to bind its target or epitope such that thedissociation constant (K_(d)) of the AB1 when coupled to the MM1 is atleast 1,000 times greater than the K_(d) of the AB2 when coupled to theMM2. In some embodiments, the coupling of each of the MM reduces theability of the corresponding AB to bind its target or epitope such thatthe dissociation constant (K_(d)) of the AB1 when coupled to the MM1 isat least 1,000 times lower than the K_(d) of the AB2 when coupled to theMM2.

In some embodiments, the coupling of each of the MM reduces the abilityof the corresponding AB to bind its target or epitope such that thedissociation constant (K_(d)) of the AB1 when coupled to the MM1 is atleast 10,000 times greater than the K_(d) of the AB2 when coupled to theMM2. In some embodiments, the coupling of each of the MM reduces theability of the corresponding AB to bind its target or epitope such thatthe dissociation constant (K_(d)) of the AB1 when coupled to the MM1 isat least 10,000 times lower than the K_(d) of the AB2 when coupled tothe MM2.

In some embodiments, each of the MM is an amino acid sequence selectedfrom a MM shown in the Examples provided herein.

In some embodiments, the protease that cleaves the first cleavablemoiety (CM1) sequence is produced by a tumor that is in proximity tocells that express the target and/or produced by a tumor that isco-localized with the target of the AB1 in the multispecific activatableantibody in a tissue, and the protease cleaves the CM1 in themultispecific activatable antibody when the multispecific activatableantibody is exposed to the protease.

In some embodiments, the multispecific activatable antibody includesmore than one cleavable moiety sequence, and the protease that cleavesat least one cleavable moiety sequence is produced by a tumor that is inproximity to cells that express the target and/or produced by a tumorthat is co-localized with the target of at least one of the AB regionsin the multispecific activatable antibody in a tissue, and the proteasecleaves the CM in the multispecific activatable antibody when themultispecific activatable antibody is exposed to the protease.

In some embodiments, each CM, e.g., CM1 and at least CM2, is positionedin the multispecific activatable antibody such that when the activatableantibody is in the uncleaved state, binding of the multispecificactivatable antibody to a target of one of the AB regions is reduced tooccur with a dissociation constant that is at least 20-fold greater thanthe dissociation constant of an unmodified AB binding to its target, andwhereas when the activatable antibody is in the cleaved state, the ABbinds its target.

In some embodiments, each CM is positioned in the multispecificactivatable antibody such that when the activatable antibody is in theuncleaved state, binding of the multispecific activatable antibody to atarget of one of the AB regions is reduced to occur with a dissociationconstant that is at least 40-fold greater than the dissociation constantof an unmodified AB binding to its target, and whereas when theactivatable antibody is in the cleaved state, the AB binds its target.

In some embodiments, each CM is positioned in the multispecificactivatable antibody such when the activatable antibody is that in theuncleaved state, binding of the multispecific activatable antibody to atarget of one of the AB regions is reduced to occur with a dissociationconstant that is at least 50-fold greater than the dissociation constantof an unmodified AB binding to its target, and whereas when theactivatable antibody is in the cleaved state, the AB binds its target.

In some embodiments, each CM is positioned in the multispecificactivatable antibody such when the activatable antibody is that in theuncleaved state, binding of the multispecific activatable antibody to atarget of one of the AB regions is reduced to occur with a dissociationconstant that is at least 100-fold greater than the dissociationconstant of an unmodified AB binding to its target, and whereas when theactivatable antibody is in the cleaved state, the AB binds its target.

In some embodiments, each CM is positioned in the multispecificactivatable antibody such that when the activatable antibody is in theuncleaved state, binding of the multispecific activatable antibody to atarget of one of the AB regions is reduced to occur with a dissociationconstant that is at least 200-fold greater than the dissociationconstant of an unmodified AB binding to its target, and whereas when theactivatable antibody is in the cleaved state, the AB binds its target.

In some embodiments, each CM in the multispecific activatable antibodyis a polypeptide of up to 15 amino acids in length.

In some embodiments, at least one CM in the multispecific activatableantibody includes the amino acid sequence LSGRSDNH (SEQ ID NO: 67). Insome embodiments, at least one cleavable moiety is selected for use witha specific protease, for example a protease that is known to be producedby a tumor that is in proximity to cells that express the target and/orproduced by a tumor that is co-localized with at least one target of themultispecific activatable antibody. For example, suitable cleavablemoieties for use in the multispecific activatable antibodies of thedisclosure are cleaved by at least a protease such as urokinase,legumain, and/or matriptase (also referred to herein as MT-SP1 orMTSP1). In some embodiments, a suitable cleavable moiety includes atleast one of the following sequences: TGRGPSWV (SEQ ID NO: 68); SARGPSRW(SEQ ID NO: 69); TARGPSFK (SEQ ID NO: 70); LSGRSDNH (SEQ ID NO: 67);GGWHTGRN (SEQ ID NO: 71); HTGRSGAL (SEQ ID NO: 72); PLTGRSGG (SEQ ID NO:73); AARGPAIH (SEQ ID NO: 74); RGPAFNPM (SEQ ID NO: 75); SSRGPAYL (SEQID NO: 76); RGPATPIM (SEQ ID NO: 77); RGPA (SEQ ID NO: 78); GGQPSGMWGW(SEQ ID NO: 79); FPRPLGITGL (SEQ ID NO: 80); VHMPLGFLGP (SEQ ID NO: 81);SPLTGRSG (SEQ ID NO: 82); SAGFSLPA (SEQ ID NO: 83); LAPLGLQRR (SEQ IDNO: 84); SGGPLGVR (SEQ ID NO: 85); and/or PLGL (SEQ ID NO: 86).

In some embodiments, each CM in the multispecific activatable antibodyis a substrate for a protease selected from the group consisting ofthose shown in Table 3. In some embodiments, the protease is selectedfrom the group consisting of uPA, legumain, matriptase, ADAM17, BMP-1,TMPRSS3, TMPRSS4, neutrophil elastase, MMP-7, MMP-9, MMP-12, MMP-13, andMMP-14. In some embodiments, the protease is a cathepsin, such as, butnot limited to, cathepsin S. In some embodiments, each CM in themultispecific activatable antibody is a substrate for a proteaseselected from the group consisting of uPA (urokinase plasminogenactivator), legumain and matriptase. In some embodiments, the proteasecomprises uPA. In some embodiments, the protease comprises legumain. Insome embodiments, the protease comprises matriptase. In someembodiments, the protease comprises a matrix metalloproteinase (MMP).

In some embodiments, at least one CM in the multispecific activatableantibody is a substrate for at least two proteases. In some embodiments,each protease is selected from the group consisting of those shown inTable 3. In some embodiments, at least one CM in the multispecificactivatable antibody is a substrate for at least two proteases, whereinone of the proteases is selected from the group consisting of uPA,legumain and matriptase and the other protease is selected from thegroup consisting of those shown in Table 3. In some embodiments, atleast one CM in the multispecific activatable antibody is a substratefor at least two proteases selected from the group consisting of uPA,legumain and matriptase.

In some embodiments, at least one of the proteases that can cleave eachCM in the multispecific activatable antibody is the same protease. Insome embodiments, the same protease that can cleave both CMs in themultispecific activatable antibody is selected from the group consistingof those shown in Table 3. In some embodiments, the same protease thatcan cleave both CMs in the multispecific activatable antibody areselected from the group consisting of uPA, legumain and matriptase.

In some embodiments, at least two of the proteases that can cleave eachCM in the multispecific activatable antibody are the same two proteases.In some embodiments, the same two proteases that can cleave both CMs inthe multispecific activatable antibody are selected from the groupconsisting of those shown in Table 3. In some embodiments, at least oneof the at least the two proteases that can cleave both CMs in themultispecific activatable antibody is selected from the group consistingof uPA, legumain and matriptase, and the other protease of the at leastthe two proteases is selected from the group consisting of those shownin Table 3. In some embodiments, the at least the two proteases that cancleave both CMs in the multispecific activatable antibody are selectedfrom the group consisting of uPA, legumain and matriptase.

In some embodiments, at least one of the proteases that can cleave CM1is different than at least one of the proteases that can cleave CM2 inthe multispecific activatable antibody. In some embodiments, at leastone of the proteases that can cleave CM1 but cannot cleave CM2 in themultispecific activatable antibody. In some embodiments, at least one ofthe proteases that can cleave CM2 but cannot cleave CM1 in themultispecific activatable antibody. In some embodiments, the at leastone protease that can cleave CM1 but cannot cleave CM2 in themultispecific activatable antibody is selected from the group consistingof those shown in Table 3. In some embodiments, the at least oneprotease that can cleave CM1 but cannot cleave CM2 in the multispecificactivatable is selected from the group consisting of uPA, legumain andmatriptase.

In some embodiments, the multispecific activatable antibody includes atleast a first CM (CM1) and a second CM (CM2). In some embodiments, CM1and CM2 are part of a single cleavable linker that joins an MM to an AB.In some embodiments, CM1 is part of a cleavable linker that joins MM1 toAB1, and CM2 is part of a separate cleavable linker that joins an MM2 toAB2. In some embodiments, a multispecific activatable antibody comprisesmore than two CMs. In some embodiments, such a multispecific activatableantibody comprises more than two CMs and more than two MMs. In someembodiments, CM1 and CM2 are each polypeptides of no more than 15 aminoacids long. In some embodiments, at least one of the first CM and thesecond CM is a polypeptide that functions as a substrate for a proteaseselected from the group consisting of those listed in Table 3. In someembodiments, at least one of the first CM and the second CM is apolypeptide that functions as a substrate for a protease selected fromthe group consisting of uPA, legumain, and matriptase. In someembodiments, the first CM is cleaved by a first cleaving agent selectedfrom the group consisting of uPA, legumain, and matriptase in a targettissue and the second CM is cleaved by a second cleaving agent in atarget tissue. In some embodiments, the other protease is selected fromthe group consisting of those shown in Table 3. In some embodiments, thefirst cleaving agent and the second cleaving agent are the same proteaseselected from the group consisting of those listed in Table 3, and thefirst CM and the second CM are different substrates for the protease. Insome embodiments, the first cleaving agent and the second cleaving agentare the same protease selected from the group consisting of uPA,legumain, and matriptase, and the first CM and the second CM aredifferent substrates for the protease. In some embodiments, the firstcleaving agent and the second cleaving agent are the same proteaseselected from the group listed in Table 3, and the first CM and thesecond CM are the same substrate. In some embodiments, the firstcleaving agent and the second cleaving agent are different proteases. Insome embodiments, the first cleaving agent and the second cleaving agentare different proteases selected from the group consisting of thoseshown in Table 3. In some embodiments, the first cleaving agent and thesecond cleaving agent are produced by a tumor that is in proximity tocells that express the target and/or produced by a tumor that isco-localized in the target tissue. In some embodiments, the first CM andthe second CM are cleaved by at least one cleaving agent in the targettissue.

In some embodiments, the multispecific activatable antibody is exposedto and cleaved by a protease such that, when the activatable antibody isin the activated or cleaved state, the activated multispecificactivatable antibody includes a light chain amino acid sequence thatincludes at least a portion of LP2 and/or CM sequence after the proteasehas cleaved the CM.

In some embodiments, the multispecific activatable antibody alsoincludes a signal peptide. In some embodiments, the signal peptide isconjugated to the multispecific activatable antibody via a spacer. Insome embodiments, the spacer is conjugated to the multispecificactivatable antibody in the absence of a signal peptide. In someembodiments, the spacer is joined directly to at least one of the MM ofthe multispecific activatable antibody.

In some embodiments, the multispecific activatable antibody in anuncleaved state comprises a spacer that is joined directly to a first MMand has the structural arrangement from N-terminus to C-terminus ofspacer-MM1-CM-AB1. In some embodiments, the spacer includes at least theamino acid sequence QGQSGQ (SEQ ID NO: 87).

In some embodiments, the serum half-life of the multispecificactivatable antibody is longer than that of the correspondingmultispecific antibody; e.g., the pK of the multispecific activatableantibody is longer than that of the corresponding multispecificantibody. In some embodiments, the serum half-life of the multispecificactivatable antibody is similar to that of the correspondingmultispecific antibody. In some embodiments, the serum half-life of themultispecific activatable antibody is at least 15 days when administeredto an organism. In some embodiments, the serum half-life of themultispecific activatable antibody is at least 12 days when administeredto an organism. In some embodiments, the serum half-life of themultispecific activatable antibody is at least 11 days when administeredto an organism. In some embodiments, the serum half-life of themultispecific activatable antibody is at least 10 days when administeredto an organism. In some embodiments, the serum half-life of themultispecific activatable antibody is at least 9 days when administeredto an organism. In some embodiments, the serum half-life of themultispecific activatable antibody is at least 8 days when administeredto an organism. In some embodiments, the serum half-life of themultispecific activatable antibody is at least 7 days when administeredto an organism. In some embodiments, the serum half-life of themultispecific activatable antibody is at least 6 days when administeredto an organism. In some embodiments, the serum half-life of themultispecific activatable antibody is at least 5 days when administeredto an organism. In some embodiments, the serum half-life of themultispecific activatable antibody is at least 4 days when administeredto an organism. In some embodiments, the serum half-life of themultispecific activatable antibody is at least 3 days when administeredto an organism. In some embodiments, the serum half-life of themultispecific activatable antibody is at least 2 days when administeredto an organism. In some embodiments, the serum half-life of themultispecific activatable antibody is at least 24 hours whenadministered to an organism. In some embodiments, the serum half-life ofthe multispecific activatable antibody is at least 20 hours whenadministered to an organism. In some embodiments, the serum half-life ofthe multispecific activatable antibody is at least 18 hours whenadministered to an organism. In some embodiments, the serum half-life ofthe multispecific activatable antibody is at least 16 hours whenadministered to an organism. In some embodiments, the serum half-life ofthe multispecific activatable antibody is at least 14 hours whenadministered to an organism. In some embodiments, the serum half-life ofthe multispecific activatable antibody is at least 12 hours whenadministered to an organism. In some embodiments, the serum half-life ofthe multispecific activatable antibody is at least 10 hours whenadministered to an organism. In some embodiments, the serum half-life ofthe multispecific activatable antibody is at least 8 hours whenadministered to an organism. In some embodiments, the serum half-life ofthe multispecific activatable antibody is at least 6 hours whenadministered to an organism. In some embodiments, the serum half-life ofthe multispecific activatable antibody is at least 4 hours whenadministered to an organism. In some embodiments, the serum half-life ofthe multispecific activatable antibody is at least 3 hours whenadministered to an organism.

The disclosure also provides compositions and methods that include amultispecific activatable antibody that includes at least a firstantibody or antibody fragment (AB1) that specifically binds a target anda second antibody or antibody fragment (AB2), where at least the firstAB in the multispecific activatable antibody is coupled to a maskingmoiety (MM1) that decreases the ability of AB1 to bind its target. Insome embodiments, each AB is coupled to a MM that decreases the abilityof its corresponding AB to each target. For example, in bispecificactivatable antibody embodiments, AB1 is coupled to a first maskingmoiety (MM1) that decreases the ability of AB1 to bind its target, andAB2 is coupled to a second masking moiety (MM2) that decreases theability of AB2 to bind its target. In some embodiments, themultispecific activatable antibody comprises more than two AB regions;in such embodiments, AB1 is coupled to a first masking moiety (MM1) thatdecreases the ability of AB1 to bind its target, AB2 is coupled to asecond masking moiety (MM2) that decreases the ability of AB2 to bindits target, AB3 is coupled to a third masking moiety (MM3) thatdecreases the ability of AB3 to bind its target, and so on for each ABin the multispecific activatable antibody.

In some embodiments, the multispecific activatable antibody furtherincludes at least one cleavable moiety (CM) that is a substrate for aprotease, where the CM links a MM to an AB. For example, in someembodiments, the multispecific activatable antibody includes at least afirst antibody or antibody fragment (AB1) that specifically binds atarget and a second antibody or antibody fragment (AB2), where at leastthe first AB in the multispecific activatable antibody is coupled via afirst cleavable moiety (CM1) to a masking moiety (MM1) that decreasesthe ability of AB1 to bind its target. In some bispecific activatableantibody embodiments, AB1 is coupled via CM1 to MM1, and AB2 is coupledvia a second cleavable moiety (CM2) to a second masking moiety (MM2)that decreases the ability of AB2 to bind its target. In someembodiments, the multispecific activatable antibody comprises more thantwo AB regions; in some of these embodiments, AB1 is coupled via CM1 toMM1, AB2 is coupled via CM2 to MM2, and AB3 is coupled via a thirdcleavable moiety (CM3) to a third masking moiety (MM3) that decreasesthe ability of AB3 to bind its target, and so on for each AB in themultispecific activatable antibody.

In some embodiments, the multispecific antibodies and/or multispecificactivatable antibodies described herein are used in conjunction with oneor more additional agents or a combination of additional agents.Suitable additional agents include current pharmaceutical and/orsurgical therapies for an intended application, such as, for example,cancer. For example, the multispecific antibodies and/or multispecificactivatable antibodies can be used in conjunction with an additionalchemotherapeutic or anti-neoplastic agent.

In some embodiments, the multispecific antibody and/or multispecificactivatable antibody and additional agent are formulated into a singletherapeutic composition, and the multispecific antibody and/ormultispecific activatable antibody and additional agent are administeredsimultaneously. In some embodiments, the multispecific antibody and/ormultispecific activatable antibody and additional agent are separatefrom each other, e.g., each is formulated into a separate therapeuticcomposition, and the multispecific antibody and/or multispecificactivatable antibody and the additional agent are administeredsimultaneously, or the multispecific antibody and/or multispecificactivatable antibody and the additional agent are administered atdifferent times during a treatment regimen. For example, themultispecific antibody and/or multispecific activatable antibody isadministered prior to the administration of the additional agent, themultispecific antibody and/or multispecific activatable antibody isadministered subsequent to the administration of the additional agent,or the multispecific antibody and/or multispecific activatable antibodyand the additional agent are administered in an alternating fashion. Asdescribed herein, the anti-multispecific antibody and/or multispecificactivatable antibody and additional agent are administered in singledoses or in multiple doses.

The disclosure also provides an isolated nucleic acid molecule encodingat least a portion of a multispecific antibody and/or multispecificactivatable antibody described herein, such as for example, at least afirst nucleic acid encoding at least a portion of the heavy chain of themultispecific antibody and/or multispecific activatable antibody and asecond nucleic acid encoding at least a portion of the light chain ofmultispecific antibody and/or multispecific the activatable antibody, aswell as vectors that include these isolated nucleic acid sequences. Thedisclosure provides methods of producing a multispecific antibody and/ormultispecific activatable antibody by culturing a cell under conditionsthat lead to expression of the multispecific antibody and/ormultispecific activatable antibody, wherein the cell comprises such anucleic acid molecule(s). In some embodiments, the cell comprises such avector.

The disclosure also provides a method of manufacturing multispecificantibodies of the disclosure and/or multispecific activatable antibodiesof the disclosure by (a) culturing a cell comprising a nucleic acidconstruct that encodes the multispecific antibody and/or multispecificactivatable antibody under conditions that lead to expression of themultispecific antibody and/or multispecific activatable, and (b)recovering the multispecific antibody and/or multispecific activatableantibody.

The present disclosure also provides methods of treating, preventing,delaying the progression of or otherwise ameliorating a symptom of oneor more pathologies or alleviating a symptom associated with suchpathologies, by administering an anti-CD3ε antibody, an activatableanti-CD3ε antibody, a multispecific antibody that specifically bindsCD3ε and/or a multispecific activatable antibody of the disclosure thatspecifically binds CD3ε to a subject in which such treatment orprevention is desired. The subject to be treated is, e.g., human orother mammal. In some embodiments, the subject is a non-human mammal,such as a non-human primate, companion animal (e.g., cat, dog, horse),farm animal, work animal, or zoo animal. In some embodiments, thesubject is a rodent.

The present disclosure also provides methods to induce target-dependentT-cell activation and killing of a target cell by administering amultispecific activatable antibody of the disclosure to a subject inwhich such induction is desired, wherein when the multispecificactivatable antibody is in the cleaved state, e.g., each masking moietyin the multispecific activatable antibody is no longer attached orotherwise associated with the corresponding AB domain, target-dependentT-cell activation and killing of the target cell occurs, and when themultispecific activatable antibody is in the uncleaved state, e.g. atleast one masking moiety of the multispecific activatable antibody isattached or otherwise associated with the corresponding AB domain,target-dependent T-cell activation and killing of the target cell isreduced or otherwise inhibited. Any of the multispecific activatableantibodies described herein are suitable for use in such methods. Thesubject to be treated is, e.g., human or other mammal. In someembodiments, the subject is a non-human mammal, such as a non-humanprimate, companion animal (e.g., cat, dog, horse), farm animal, workanimal, or zoo animal. In some embodiments, the subject is a rodent.

An anti-CD3ε antibody, an activatable anti-CD3ε antibody, amultispecific antibody that specifically binds CD3ε and/or amultispecific activatable antibody of the disclosure that specificallybinds CD3ε used in any of the embodiments of these methods and uses canbe administered at any stage of the disease. For example, such anantibody, an activatable antibody, a multispecific antibody and/or amultispecific activatable antibody can be administered to a patientsuffering cancer of any stage, from early to metastatic. The termssubject and patient are used interchangeably herein.

An anti-CD3ε antibody, an activatable anti-CD3ε antibody, amultispecific antibody that specifically binds CD3ε and/or amultispecific activatable antibody of the disclosure that specificallybinds CD3ε used in any of the embodiments of these methods and uses canbe used in a treatment regimen comprising neoadjuvant therapy.

An anti-CD3ε antibody, an activatable anti-CD3ε antibody, amultispecific antibody that specifically binds CD3ε and/or amultispecific activatable antibody of the disclosure that specificallybinds CD3ε used in any of the embodiments of these methods and uses canbe administered either alone or in combination with one or moreadditional agents, including small molecule inhibitors, otherantibody-based therapies, polypeptide or peptide-based therapies,nucleic acid-based therapies and/or other biologics. In someembodiments, an antibody, an activatable antibody, a multispecificantibody, and/or a multispecific activatable antibody is administered incombination with one or more additional agents such as, by way ofnon-limiting example, a chemotherapeutic agent, such as an alkylatingagent, an anti-metabolite, an anti-microtubule agent, a topoisomeraseinhibitor, a cytotoxic antibiotic, and any other nucleic acid damagingagent. In some embodiments, the additional agent is a taxane, such aspaclitaxel (e.g., Abraxane®). In some embodiments, the additional agentis an anti-metabolite, such as gemcitabine. In some embodiments, theadditional agent is an alkylating agent, such as platinum-basedchemotherapy, such as carboplatin or cisplatin. In some embodiments, theadditional agent is a targeted agent, such as a kinase inhibitor, e.g.,sorafenib or erlotinib. In some embodiments, the additional agent is atargeted agent, such as another antibody, e.g., a monoclonal antibody(e.g., bevacizumab), a bispecific antibody, or a multispecific antibody.In some embodiments, the additional agent is a proteosome inhibitor,such as bortezomib or carfilzomib. In some embodiments, the additionalagent is an immune modulating agent, such as lenolidominde or IL-2. Insome embodiments, the additional agent is radiation. In someembodiments, the additional agent is an agent considered standard ofcare by those skilled in the art. In some embodiments, the additionalagent is a chemotherapeutic agent well known to those skilled in theart. In some embodiments, the antibody, activatable antibody,multispecific antibody, and/or multispecific activatable antibody, andthe additional agent(s) are formulated in a single composition. In someembodiments, the antibody, activatable antibody, multispecific antibody,and/or multispecific activatable antibody, and the additional agent(s)are administered as two or more separate compositions. In someembodiments, the antibody, activatable antibody, multispecific antibody,and/or multispecific activatable antibody, and the additional agent(s)are administered simultaneously. In some embodiments, the antibody,activatable antibody, multispecific antibody, and/or multispecificactivatable antibody, and the additional agent(s) are administeredsequentially.

In some embodiments, the additional agent(s) is a chemotherapeuticagent, such as a chemotherapeutic agent selected from the groupconsisting of docetaxel, paclitaxel, abraxane (i.e., albumin-conjugatedpaclitaxel), doxorubicin, oxaliplatin, carboplatin, cisplatin,irinotecan, and gemcitabine.

In some embodiments, the additional agent(s) is a checkpoint inhibitor,a kinase inhibitor, an agent targeting inhibitors in the tumormicroenvironment, and/or a T cell or NK agonist. In some embodiments,the additional agent(s) is radiation therapy, alone or in combinationwith another additional agent(s) such as a chemotherapeutic oranti-neoplastic agent. In some embodiments, the additional agent(s) is avaccine, an oncovirus, and/or a DC-activating agent such as, by way ofnon-limiting example, a toll-like receptor (TLR) agonist and/or α-CD40.In some embodiments, the additional agent(s) is a tumor-targetedantibody designed to kill the tumor via ADCC or via direct conjugationto a toxin (e.g., an antibody drug conjugate (ADC).

In some embodiments, the checkpoint inhibitor is an inhibitor of atarget selected from the group consisting of CTLA-4, LAG-3, PD-1, PDL1,TIGIT, TIM-3, B7H4, and Vista. In some embodiments, the kinase inhibitoris selected from the group consisting of B-RAFi, MEKi, and Btkinhibitors, such as ibrutinib. In some embodiments, the kinase inhibitoris crizotinib. In some embodiments, the tumor microenvironment inhibitoris selected from the group consisting of an IDO inhibitor, an α-CSF1Rinhibitor, an α-CCR4 inhibitor, a TGF-beta, a myeloid-derived suppressorcell, or a T-regulatory cell. In some embodiments, the agonist isselected from the group consisting of Ox40, GITR, CD137, ICOS, CD27, andHVEM. In some embodiments, the checkpoint inhibitor is an antibody thatbinds a target selected from CTLA-4, PD-1, and/or PD-L1. In someembodiments, the checkpoint inhibitor is an anti-CTLA4 antibody, ananti-PD-1 antibody, and an anti-PD-L1 antibody, and/or combinationsthereof. In some embodiments, the checkpoint inhibitor is an anti-CTLA4antibody such as, e.g., Yervoy™. In some embodiments, the checkpointinhibitor is an anti-PD-1 antibody such as, e.g., Opdivo™ and/orKeytruda™.

In some embodiments, the inhibitor is a CTLA-4 inhibitor. In someembodiments, the inhibitor is a LAG-3 inhibitor. In some embodiments,the inhibitor is a PD-1 inhibitor. In some embodiments, the inhibitor isa PDL1 inhibitor. In some embodiments, the inhibitor is a TIGITinhibitor. In some embodiments, the inhibitor is a TIM-3 inhibitor. Insome embodiments, the inhibitor is a B7H4 inhibitor. In someembodiments, the inhibitor is a Vista inhibitor. In some embodiments,the inhibitor is a B-RAFi inhibitor. In some embodiments, the inhibitoris a MEKi inhibitor. In some embodiments, the inhibitor is a Btkinhibitor. In some embodiments, the inhibitor is ibrutinib. In someembodiments, the inhibitor is crizotinib. In some embodiments, theinhibitor is an IDO inhibitor. In some embodiments, the inhibitor is anα-CSF1R inhibitor. In some embodiments, the inhibitor is an α-CCR4inhibitor. In some embodiments, the inhibitor is a TGF-beta. In someembodiments, the inhibitor is a myeloid-derived suppressor cell. In someembodiments, the inhibitor is a T-regulatory cell.

In some embodiments, the agonist is Ox40. In some embodiments, theagonist is GITR. In some embodiments, the agonist is CD137. In someembodiments, the agonist is ICOS. In some embodiments, the agonist isCD27. In some embodiments, the agonist is HVEM.

In some embodiments, the additional agent is another antibody orantigen-binding fragment thereof, another conjugated antibody orantigen-binding fragment thereof, another activatable antibody orantigen-binding fragment thereof and/or another conjugated activatableantibody or antigen-binding fragment thereof. In some embodiments theadditional agent is another antibody or antigen-binding fragmentthereof, another conjugated antibody or antigen-binding fragmentthereof, another activatable antibody or antigen-binding fragmentthereof and/or another conjugated activatable antibody orantigen-binding fragment thereof against the same target as the firstantibody or antigen-binding fragment thereof, the first conjugatedantibody or antigen-binding fragment thereof, activatable antibody orantigen-binding fragment thereof and/or a conjugated activatableantibody or antigen-binding fragment thereof, e.g., against PDL1. Insome embodiments the additional agent is another antibody orantigen-binding fragment thereof, another conjugated antibody orantigen-binding fragment thereof, another activatable antibody orantigen-binding fragment thereof and/or another conjugated activatableantibody or antigen-binding fragment thereof against a target differentthan the target of the first antibody or antigen-binding fragmentthereof, the first conjugated antibody or antigen-binding fragmentthereof, activatable antibody or antigen-binding fragment thereof and/ora conjugated activatable antibody or antigen-binding fragment thereof.

In some embodiments, the anti-CD3ε antibody and/or activatable antibodyis administered during and/or after treatment in combination with one ormore additional agents such as, for example, a chemotherapeutic agent,an anti-inflammatory agent, and/or a an immunosuppressive agent. In someembodiments, the anti-CD3ε antibody and/or activatable antibody and theadditional agent are formulated into a single therapeutic composition,and the anti-CD3ε antibody and/or activatable antibody and additionalagent are administered simultaneously. Alternatively, the anti-CD3εantibody and/or activatable antibody and additional agent are separatefrom each other, e.g., each is formulated into a separate therapeuticcomposition, and the anti-CD3ε antibody and/or activatable antibody andthe additional agent are administered simultaneously, or the anti-CD3εantibody and/or activatable antibody and the additional agent areadministered at different times during a treatment regimen. For example,the anti-CD3ε antibody and/or activatable antibody is administered priorto the administration of the additional agent, the anti-CD3ε antibodyand/or activatable antibody is administered subsequent to theadministration of the additional agent, or the anti-CD3ε antibody and/oractivatable antibody and the additional agent are administered in analternating fashion. As described herein, the anti-CD3ε antibody and/oractivatable antibody and additional agent are administered in singledoses or in multiple doses.

In some embodiments, the anti-CD3ε antibody and/or activatable antibodyand the additional agent(s) are administered simultaneously. Forexample, the anti-CD3ε antibody and/or activatable antibody and theadditional agent(s) can be formulated in a single composition oradministered as two or more separate compositions. In some embodiments,the anti-CD3ε antibody and/or activatable antibody and the additionalagent(s) are administered sequentially, or the anti-CD3ε antibody and/oractivatable antibody and the additional agent are administered atdifferent times during a treatment regimen.

The disclosure also provides methods and kits for using the anti-CD3εantibody, activatable anti-CD3ε antibody, multispecific antibody thatspecifically binds CD3ε and/or multispecific activatable antibody of thedisclosure that specifically binds CD3ε in a variety of diagnosticand/or prophylactic indications. For example, the disclosure providesmethods and kits for detecting presence or absence of a cleaving agentand a target of interest in a subject or a sample by (i) contacting asubject or sample with an activatable antibody or a multispecificactivatable antibody that includes at least a first masking moiety(MM1), a first cleavable moiety (CM1) that is cleaved by the cleavingagent, and at least a first antigen binding domain or fragment thereof(AB1) that specifically binds the target of interest, (a) wherein theMM1 is a peptide that inhibits binding of the AB1 to the target, andwherein the MM1 does not have an amino acid sequence of a naturallyoccurring binding partner of the AB1 and is not a modified form of anatural binding partner of the AB1; and (b) wherein, in an uncleaved,non-activated state, the MM1 interferes with specific binding of the AB1to the target, and in a cleaved, activated state the MM1 does notinterfere or compete with specific binding of the AB1 to the target; and(ii) measuring a level of activated antibody or activated multispecificactivatable antibody in the subject or sample, wherein a detectablelevel of activated antibody or activated multispecific activatableantibody in the subject or sample indicates that the cleaving agent andthe target are present in the subject or sample and wherein nodetectable level of activated antibody or activated multispecificactivatable antibody in the subject or sample indicates that thecleaving agent, the target or both the cleaving agent and the target areabsent and/or not sufficiently present in the subject or sample.

In some embodiments, the activatable antibody or activatablemultispecific activatable antibody is an activatable antibody or anactivatable multispecific activatable antibody to which a therapeuticagent is conjugated. In some embodiments, the activatable antibody oractivatable multispecific activatable antibody is not conjugated to anagent. In some embodiments, the activatable antibody or activatablemultispecific activatable antibody comprises a detectable label. In someembodiments, the detectable label is positioned on the AB1. In someembodiments, measuring the level of activatable antibody ormultispecific activatable antibody in the subject or sample isaccomplished using a secondary reagent that specifically binds to theactivatable antibody or activatable multispecific activatable antibody,wherein the reagent comprises a detectable label. In some embodiments,the secondary reagent is an antibody comprising a detectable label.

In some embodiments of these methods and kits, the activatable antibodyor multispecific activatable antibody includes a detectable label. Insome embodiments of these methods and kits, the detectable labelincludes an imaging agent, a contrasting agent, an enzyme, a fluorescentlabel, a chromophore, a dye, one or more metal ions, or a ligand-basedlabel. In some embodiments of these methods and kits, the imaging agentcomprises a radioisotope. In some embodiments of these methods and kits,the radioisotope is indium or technetium. In some embodiments of thesemethods and kits, the contrasting agent comprises iodine, gadolinium oriron oxide. In some embodiments of these methods and kits, the enzymecomprises horseradish peroxidase, alkaline phosphatase, orβ-galactosidase. In some embodiments of these methods and kits, thefluorescent label comprises yellow fluorescent protein (YFP), cyanfluorescent protein (CFP), green fluorescent protein (GFP), modified redfluorescent protein (mRFP), red fluorescent protein tdimer2 (RFPtdimer2), HCRED, or a europium derivative. In some embodiments of thesemethods and kits, the luminescent label comprises an N-methylacrydiumderivative. In some embodiments of these methods, the label comprises anAlexa Fluor® label, such as Alex Fluor® 680 or Alexa Fluor® 750. In someembodiments of these methods and kits, the ligand-based label comprisesbiotin, avidin, streptavidin or one or more haptens.

In some embodiments of these methods and kits, the subject is a mammal.In some embodiments of these methods and kits, the subject is a human.In some embodiments, the subject is a non-human mammal, such as anon-human primate, companion animal (e.g., cat, dog, horse), farmanimal, work animal, or zoo animal. In some embodiments, the subject isa rodent.

In some embodiments of these methods, the method is an in vivo method.In some embodiments of these methods, the method is an in situ method.In some embodiments of these methods, the method is an ex vivo method.In some embodiments of these methods, the method is an in vitro method.

In some embodiments of the methods and kits, the method or kit is usedto identify or otherwise refine a patient population suitable fortreatment with a multispecific activatable antibody of the disclosure.For example, patients that test positive for both the target and aprotease that cleaves the substrate in the first cleavable moiety (CM1)of the multispecific activatable antibody being tested in these methodsare identified as suitable candidates for treatment with such anactivatable antibody or a multispecific activatable antibody comprisingsuch a CM1. Likewise, patients that test negative for both the targetand the protease that cleaves the substrate in the CM1 in theactivatable antibody or multispecific activatable antibody being testedusing these methods might be identified as suitable candidates foranother form of therapy.

In some embodiments, a method or kit is used to identify or otherwiserefine a patient population suitable for treatment with an activatableantibody and/or a multispecific activatable antibody of the disclosure,followed by treatment by administering that activatable antibody and/ormultispecific activatable antibody to a subject in need thereof. Forexample, patients that test positive for both the target and a proteasethat cleaves the substrate in the first cleavable moiety (CM1) of theactivatable antibody and/or multispecific activatable antibody beingtested in these methods are identified as suitable candidates fortreatment with an activatable antibody and/or a multispecificactivatable antibody comprising such a CM1, and the patient is thenadministered a therapeutically effective amount of the activatableantibody and/or the multispecific activatable antibody that was tested.Likewise, patients that test negative for either or both of the targetand the protease that cleaves the substrate in the CM1 in theactivatable antibody and/or multispecific activatable antibody beingtested using these methods might be identified as suitable candidatesfor another form of therapy.

In some embodiments, such patients can be tested with other activatableantibodies, conjugated activatable antibodies or multispecificantibodies and/or multispecific activatable antibodies and/or conjugatedmultispecific activatable antibodies until a suitable activatableantibody or conjugated activatable antibody or multispecific activatableantibody and/or conjugated multispecific activatable antibody fortreatment is identified, e.g., an activatable antibody or a conjugatedactivatable antibody or a multispecific activatable antibody and/orconjugated multispecific activatable antibody comprising a CM that iscleaved by the patient at the site of disease. In some embodiments, thepatient is then administered a therapeutically effective amount of theactivatable antibody or conjugated activatable antibody or multispecificactivatable antibody and/or conjugated multispecific activatableantibody for which the patient tested positive.

Pharmaceutical compositions according to the disclosure can include amultispecific antibody and/or a multispecific activatable antibody ofthe disclosure and a carrier. These pharmaceutical compositions can beincluded in kits, such as, for example, diagnostic kits.

One skilled in the art will appreciate that the antibodies of thedisclosure have a variety of uses. For example, the proteins of thedisclosure are used as therapeutic agents for a variety of disorders.The antibodies of the disclosure are also used as reagents in diagnostickits or as diagnostic tools, or these antibodies can be used incompetition assays to generate therapeutic reagents.

BRIEF DESCRIPTION OF DRAWINGS

FIG. 1 is an illustration depicting generic mask selection populationnomenclature used herein.

FIGS. 2A and 2B are a series of graphs depicting the ability of scFvformats of SP34 to bind to cynomolgus CD3ε. FIG. 1A depictsSP34scFv(LvHv)-Fc and SP34scFv(HvLv)-Fc binding as a function of Fcfusion protein concentration. FIG. 2B depicts mouse SP34-2 IgG1 bindingas a function of concentration.

FIGS. 3A-3D are a series of graphs depicting the ability of maskingmoieties of the disclosure to reduce the ability of activatableanti-CD3ε antibodies comprising such masking moieties to bind to CD3ε onJurkat T cells. Each activatable antibody is labeled by its maskingmoiety name. Antibody SP34scFv(LvHv)-Fc is identified by LvHv.

FIGS. 4A, 4B, and 4C are a series of illustrations depicting variousantibodies and activatable antibodies of the disclosure. FIG. 4A depictscomponent pieces for scFv-Fc antibodies. FIG. 4B depicts scFv(LvHv)-Fcand scFv(HvLv)-Fc antibodies of the disclosure. FIG. 4C depictsactivatable antibodies comprising scFv(LvHv)-Fc or scFv(HvLv)-Fcantibodies of the disclosure.

FIGS. 5A, 5B, 5C, and 5D are a series of illustrations depictingmultispecific antibodies and multispecific activatable antibodies of thedisclosure. FIG. 5A depicts a multispecific antibody of the disclosurethat comprises an Ig antibody and a scFv. FIG. 5B depicts amultispecific activatable antibody of the disclosure in which theprimary antibody site is masked. FIG. 5C depicts a multispecificactivatable antibody of the disclosure in which the secondary scFv siteis masked. FIG. 5D depicts a multispecific activatable antibody of thedisclosure in which the with primary antibody and secondary scFv sitesare masked.

FIGS. 6A, 6B, 6C, 6D, 6E, and 6F are a series of illustrations depictingvarious embodiments of multispecific activatable antibodies of thedisclosure.

FIGS. 7A, 7B, 7C, 7D, 7E, 7F, 7G, 7H, 7I, and 7J are a series ofschematic diagrams of a selected set of the possible permutations ofmultispecific antibodies of the disclosure.

FIGS. 8A, 8B, 8C, 8D, 8E, 8F, 8G, 8H, 81, and 8J are a series ofschematic diagrams of a selected set of the possible permutations ofmultispecific activatable antibodies of the disclosure. In particular,this figure shows multispecific activatable antibodies in which theprimary antigen binding site is masked (i.e., activatable) and theadditional antigen-binding domain(s) is not masked.

FIGS. 9A, 9B, 9C, 9D, 9E, 9F, 9G, 9H, and 9J are a series of schematicdiagrams of an array of multispecific activatable antibodies in whichall antigen-binding domains are masked.

FIGS. 10A, 10B, 10C, 10D, 10E, 10F, 10G, 10H, 10I, and 10J are a seriesof schematic diagrams of an array of multispecific activatableantibodies in which the secondary antigen-binding domain is masked andthe additional antigen-binding domain(s) is not masked.

FIGS. 11A, 11B, 11C, 11D, 11E, 11F, 11G, 11H, 11I, and 11J are a seriesof schematic diagrams of an array of multispecific activatableantibodies in which the majority, but not all of the antigen-bindingdomains, are masked and at least one additional antigen-bindingdomain(s) is not masked.

FIGS. 12A, 12B, 12C, and 12D are a series of schematic diagrams of anarray of multispecific activatable antibodies in which the primaryantigen-binding domain and another antigen-binding domain are masked,and the remaining antigen-binding domain(s) is not masked.

FIGS. 13A and 13B are a series of graphs depicting SP34 scFv-Fc bindingto CD3ε positive Jurkat cells. FIG. 13A is a graph depicting the abilityof both orientations of the SP34scFv-Fc to bind to CD3ε Jurkat cells.FIG. 13B is a graph depicting the binding of SP34-2 IgG. All formats ofSP34 bind with similar EC₅₀ values.

FIGS. 14A and 14B are a series of graphs depicting the ability ofmasking moieties of the disclosure to reduce the ability of activatableanti-CD3ε antibodies comprising such masking moieties to bind to CD3ε onJurkat T cells.

FIGS. 15A, 15B, 15C, and 15D are a series of graphs depicting that theCD3ε masking peptides shift the EC₅₀ of CD3ε binding in an EGFR masked,multispecific activatable antibody that utilize two versions of an SP34scFv while retaining effective EGFR masking.

FIGS. 16A and 16B are a series of graphs depicting that the cytotoxicityEC₅₀ of CD3ε and EGFR masked multispecific, activatable antibodies areshifted relative to an unmasked control. This was evident with twoversions of the SP34 scFv, namely the scFv in CI023 and CI024 versus thescFv in CI011 and CI010.

FIGS. 17A and 17B are a series of graphs depicting that in the contextof a multi-specific, activatable antibody, the EGFR binding EC₅₀ isunaffected by the presence or absence of both CD3ε masks; likewise, CD3εEC₅₀ is unaffected by the presence or absence of the EGFR mask.

FIGS. 18A and 18B are a series of graphs depicting that the largestshift in cytotoxicity EC₅₀ is obtained by masking both EGFR and CD3binding. In addition, the cytotoxicity of the multispecific antibody andmultispecific activatable antibodies is EGFR dependent.

FIG. 19 is a graph depicting that the largest shift in activation EC₅₀is obtained by masking both EGFR and CD3 binding.

FIGS. 20A and 20B are a series of graphs depicting that CD3 maskedmultispecific antibodies show negligible target independent activationas evidenced by CD69+ frequency analysis shown in FIG. 20A and CD69 MeanFluorescence Intensity analysis shown in FIG. 20B.

FIGS. 21A, 21B, 21C, and 21D are a series of graphs depicting that uPAactivation restores binding of the multispecific, activatable CI011(M-EGFR/M-hCD3) antibody. Cell killing of all multispecific, activatableantibodies is restored by uPA activation.

FIG. 22 is a graph depicting the ability of dually masked multispecific,activatable antibodies that contain different substrates to shift EC₅₀cytotoxicity.

FIG. 23 is a graph depicting that bispecific antibody CI005 at 0.1 mg/kgdosed twice over 7 days and CI059 (BiTE) at 0.5 mg/kg/day for 8 daysprevented the growth of HT-29Luc2 xenograft tumors.

FIG. 24 is a graph depicting that CI048 antibody limited establishmentof HT-29Luc2 xenograft tumors and CI011 inhibited the growth ofHT-29Luc2 xenograft tumors. The difference in effect between PBS andCI011 is significant (p<0.05).

FIG. 25 is a graph depicting that a dose-response for the CI048bispecific antibody for elimination of HT-29Luc2 xenograft tumors.

FIG. 26 is a graph depicting that the CI048 bispecific antibody at 0.3mg/kg eliminated HT-29Luc2 xenograft tumors and the CI040 bispecificactivatable antibody at 0.1 mg/kg limited the growth of HT-29Luc2xenograft tumors.

FIG. 27 is a graph depicting that the CI040 bispecific activatableantibody at 1.0 mg/kg and the CI048 bispecific antibody at 0.3 mg/kgeliminated HT-29Luc2 xenograft tumors.

FIG. 28 is a graph depicting that the CI011 bispecific activatableantibody at 1.0 mg/kg significantly inhibited the HT-29Luc2 xenografttumor growth (p<0.05).

FIG. 29 is a graph depicting dose-response plots for CI048, CI011 andCI048 for serum alanine aminotransferase (ALT) concentration 48 hourspost-dose.

FIG. 30 shows that mouse CD3ε masks incorporated into an EGFR masked,multispecific, activatable antibody shift the EC50 of binding to mouseCD3 while retaining effective EGFR masking.

FIG. 31 demonstrate that killing of IL6R+Molp-8 cells is maximallyattenuated by masking both IL6R and CD3 binding.

DETAILED DESCRIPTION

The present disclosure provides antibodies, activatable antibodies,multispecific antibodies and/or multispecific activatable antibodiesthat bind at least the epsilon chain of CD3 (CD3ε). As used herein, anactivatable antibody is an antibody that includes a masking moiety (MM)linked to the antigen- or epitope-binding domain of the antibody suchthat coupling of the MM reduces the ability of the antigen- orepitope-binding domain to bind its target. As used herein, amultispecific antibody is an antibody that recognizes two or moredifferent antigens or epitopes, and a multispecific activatable antibodyis a multispecific antibody that includes at least one masking moiety(MM) linked to at least one antigen- or epitope-binding domain of themultispecific antibody such that coupling of the MM reduces the abilityof the antigen- or epitope-binding domain to bind its target. Theactivatable antibodies and/or activatable multispecific antibodiesprovided herein are stable in circulation, activated at intended sitesof therapy and/or diagnosis but not in normal, i.e., healthy tissue,and, when activated, exhibit binding to a target that is at leastcomparable to the corresponding, unmodified antibody or multispecificantibody.

The antibodies, activated activatable antibodies, multispecificantibodies and/or activated multispecific activatable antibodies of thepresent disclosure bind to CD3ε with a binding constant (K_(d)) of ≦1μM, for example, in some embodiments, ≦100 nM, ≦10 nM, or ≦1 nM.

The anti-CD3ε antibodies, activatable antibodies, multispecificantibodies and/or multispecific activatable antibodies of the disclosureserve to activate T cells via engagement of CD3ε on the T cells. Thatis, such antibodies agonize, stimulate, activate, and/or augmentCD3-mediated T cell activation. Biological activities of CD3 include,for example, T cell activation and other signaling through interactionbetween CD3 and the antigen-binding subunits of the T-Cell Receptor(TCR). For example, the anti-CD3ε antibodies, activated activatableantibodies, multispecific antibodies and/or activated multispecificactivatable antibodies completely or partially activate T cells viaengagement of CD3ε on T cells by partially or completely modulating,e.g., agonizing, stimulating, activating or otherwise augmentingCD3-mediated T cell activation.

Non-limiting examples of multispecific antibodies include bispecificantibodies, trispecific antibodies, tetraspecific antibodies, and othermultispecific antibodies. Multispecific antibodies provided herein arealso multivalent; as used herein, multivalency refers to the totalnumber of binding sites on the antibody, regardless of whether thebinding sites recognize the same or different antigens or epitopes.Non-limiting examples of multispecific activatable antibodies includebispecific activatable antibodies, trispecific activatable antibodies,tetraspecific activatable antibodies, and other multispecificactivatable antibodies. Multispecific activatable antibodies providedherein are also multivalent.

In some embodiments, the multispecific antibodies or fragments thereofand/or multispecific activatable antibodies or fragments thereof aredesigned to engage T cells and/or other immune effector cells.Multispecific activatable antibodies or fragments thereof that engage Tcells are also referred to herein as T-cell engaging multispecificantibodies or fragments thereof and/or T-cell engaging multispecificactivatable antibodies or fragments thereof. Multispecific activatableantibodies or fragments thereof that engage immune effector cells arealso referred to herein as immune effector cell engaging multispecificantibodies or fragments thereof and/or immune effector cell engagingmultispecific activatable antibodies or fragments thereof. In someembodiments, the multispecific antibodies or fragments thereof and/ormultispecific activatable antibodies or fragments thereof are designedto bind or otherwise interact with more than one target and/or more thanone epitope, also referred to herein as multi-antigen targetingantibodies or fragments thereof and/or multi-antigen targetingactivatable antibodies or fragments thereof.

In some embodiments, a multispecific antibody or fragment thereofincludes an IgG domain and a scFv domain. In some embodiments, amultispecific antibody or fragment thereof includes an IgG variabledomain and a scFv domain. In some embodiments, one antibody domain of amultispecific antibody or fragment thereof has specificity for a targetantigen and another antibody domain has specificity for a T-cell surfaceantigen. In some embodiments, one antibody domain of a multispecificantibody or fragment thereof has specificity for a target antigen andanother antibody domain has specificity for another target antigen. Insome embodiments, one antibody domain of a multispecific antibody orfragment thereof has specificity for an epitope of a target antigen andanother antibody domain has specificity for another epitope of the sametarget antigen.

In some embodiments, the multispecific activatable antibodies areengineered to include a masking moiety (MM) that is coupled to anantibody or antigen-binding fragment thereof (AB) via a non-cleavablelinker. For example, in some embodiments, the multispecific activatableantibody is a T-cell engaging multispecific activatable antibody thatincludes a targeting antibody or antigen-binding fragment thereof and aT-cell engaging antibody or antigen-binding portion thereof, where theT-cell engaging antibody or antigen-binding fragment thereof includes afirst antibody or antigen-binding fragment thereof (AB1) that binds afirst, T-cell engaging target, where the AB1 is attached vianon-cleavable linker to a masking moiety (MM1) such that coupling of theMM reduces the ability of the AB1 to bind the first target, and thetargeting antibody or antigen-binding fragment thereof includes a secondantibody or antigen-binding fragment thereof (AB2) that binds a secondtarget, where the AB2 is attached via a cleavable linker to a maskingmoiety (MM2) such that coupling of the MM reduces the ability of the AB2to bind the second target.

In some embodiments, the multispecific activatable antibody is a T-cellengaging multispecific activatable antibody that includes a targetingantibody or antigen-binding fragment thereof and a T-cell engagingantibody or antigen-binding portion thereof, where the T-cell engagingantibody or antigen-binding fragment thereof includes a first antibodyor antigen-binding fragment thereof (AB1) that binds a first, T-cellengaging target, where the AB1 is attached via non-cleavable linker to amasking moiety (MM1) such that coupling of the MM reduces the ability ofthe AB1 to bind the first target, and the targeting antibody orantigen-binding fragment thereof is unmasked.

DEFINITIONS

Unless otherwise defined, scientific and technical terms used inconnection with the present disclosure shall have the meanings that arecommonly understood by those of ordinary skill in the art. The term “a”entity or “an” entity refers to one or more of that entity. For example,a compound refers to one or more compounds. As such, the terms “a”,“an”, “one or more” and “at least one” can be used interchangeably.Further, unless otherwise required by context, singular terms shallinclude pluralities and plural terms shall include the singular.Generally, nomenclatures utilized in connection with, and techniques of,cell and tissue culture, molecular biology, and protein and oligo- orpolynucleotide chemistry and hybridization described herein are thosewell-known and commonly used in the art. Standard techniques are usedfor recombinant DNA, oligonucleotide synthesis, and tissue culture andtransformation (e.g., electroporation, lipofection). Enzymatic reactionsand purification techniques are performed according to manufacturer'sspecifications or as commonly accomplished in the art or as describedherein. The foregoing techniques and procedures are generally performedaccording to conventional methods well known in the art and as describedin various general and more specific references that are cited anddiscussed throughout the present specification. See e.g., Sambrook etal. Molecular Cloning: A Laboratory Manual (2d ed., Cold Spring HarborLaboratory Press, Cold Spring Harbor, N.Y. (1989)). The nomenclaturesutilized in connection with, and the laboratory procedures andtechniques of, analytical chemistry, synthetic organic chemistry, andmedicinal and pharmaceutical chemistry described herein are thosewell-known and commonly used in the art. Standard techniques are usedfor chemical syntheses, chemical analyses, pharmaceutical preparation,formulation, and delivery, and treatment of patients.

As utilized in accordance with the present disclosure, the followingterms, unless otherwise indicated, shall be understood to have thefollowing meanings:

As used herein, the term “antibody” refers to immunoglobulin moleculesand antigen-binding portions of immunoglobulin (Ig) molecules, i.e.,molecules that contain an antigen binding site that specifically binds(immunoreacts with) an antigen. By “specifically bind” or “immunoreactswith” or “immunospecifically bind” is meant that the antibody reactswith one or more antigenic determinants of the desired antigen and doesnot react with other polypeptides or binds at much lower affinity(K_(d)>10⁻⁶). Antibodies include, but are not limited to, polyclonal,monoclonal, chimeric, fully human, domain antibody, single chain, Fab,and F(ab′)₂ fragments, scFvs, and an Fab expression library.

The basic antibody structural unit is known to comprise a tetramer. Eachtetramer is composed of two identical pairs of polypeptide chains, eachpair having one “light” (about 25 kDa) and one “heavy” chain (about50-70 kDa). The amino-terminal portion of each chain includes a variableregion of about 100 to 110 or more amino acids primarily responsible forantigen recognition. The carboxy-terminal portion of each chain definesa constant region primarily responsible for effector function. Ingeneral, antibody molecules obtained from humans relate to any of theclasses IgG, IgM, IgA, IgE and IgD, which differ from one another by thenature of the heavy chain present in the molecule. Certain classes havesubclasses as well, such as IgG₁, IgG₂, IgG₃, IgG₄, and others.Furthermore, in humans, the light chain may be a kappa chain or a lambdachain.

The term “monoclonal antibody” (mAb) or “monoclonal antibodycomposition”, as used herein, refers to a population of antibodymolecules that contain only one molecular species of antibody moleculeconsisting of a unique light chain gene product and a unique heavy chaingene product. In particular, the complementarity determining regions(CDRs) of the monoclonal antibody are identical in all the molecules ofthe population. MAbs contain an antigen binding site capable ofimmunoreacting with a particular epitope of the antigen characterized bya unique binding affinity for it.

The term “antigen-binding site” or “binding portion” refers to the partof the immunoglobulin molecule that participates in antigen binding. Theantigen binding site is formed by amino acid residues of the N-terminalvariable (“V”) regions of the heavy (“H”) and light (“L”) chains. Threehighly divergent stretches within the V regions of the heavy and lightchains, referred to as “hypervariable regions,” are interposed betweenmore conserved flanking stretches known as “framework regions,” or“FRs”. Thus, the term “FR” refers to amino acid sequences that arenaturally found between, and adjacent to, hypervariable regions inimmunoglobulins. In an antibody molecule, the three hypervariableregions of a light chain and the three hypervariable regions of a heavychain are disposed relative to each other in three dimensional space toform an antigen-binding surface. The antigen-binding surface iscomplementary to the three-dimensional surface of a bound antigen, andthe three hypervariable regions of each of the heavy and light chainsare referred to as “complementarity-determining regions,” or “CDRs.” Theassignment of amino acids to each domain is in accordance with thedefinitions of Kabat Sequences of Proteins of Immunological Interest(National Institutes of Health, Bethesda, Md. (1987 and 1991)), orChothia & Lesk J. Mol. Biol. 196:901-917 (1987), Chothia et al. Nature342:878-883 (1989).

As used herein, the term “epitope” includes any protein determinantcapable of specific binding to an immunoglobulin, an scFv, or a T-cellreceptor. The term “epitope” includes any protein determinant capable ofspecific binding to an immunoglobulin or T-cell receptor. Epitopicdeterminants usually consist of chemically active surface groupings ofmolecules such as amino acids or sugar side chains and usually havespecific three dimensional structural characteristics, as well asspecific charge characteristics. For example, antibodies may be raisedagainst N-terminal or C-terminal peptides of a polypeptide. An antibodyis said to specifically bind an antigen when the dissociation constantis ≦1 μM; for example, in some embodiments ≦100 nM and in someembodiments, ≦10 nM.

As used herein, the terms “specific binding,” “immunological binding,”and “immunological binding properties” refer to the non-covalentinteractions of the type that occur between an immunoglobulin moleculeand an antigen for which the immunoglobulin is specific. The strength,or affinity of immunological binding interactions can be expressed interms of the dissociation constant (KO of the interaction, wherein asmaller K_(d) represents a greater affinity. Immunological bindingproperties of selected polypeptides can be quantified using methods wellknown in the art. One such method entails measuring the rates ofantigen-binding site/antigen complex formation and dissociation, whereinthose rates depend on the concentrations of the complex partners, theaffinity of the interaction, and geometric parameters that equallyinfluence the rate in both directions. Thus, both the “on rate constant”(K_(on)) and the “off rate constant” (K_(off)) can be determined bycalculation of the concentrations and the actual rates of associationand dissociation. (See Nature 361:186-87 (1993)). The ratio ofK_(off)/K_(on) enables the cancellation of all parameters not related toaffinity, and is equal to the dissociation constant K_(d). (See,generally, Davies et al. (1990) Annual Rev Biochem 59:439-473). Anantibody of the present disclosure is said to specifically bind to EGFR,when the binding constant (K_(d)) is ≦1 μM, for example in someembodiments ≦100 nM, in some embodiments ≦10 nM, and in some embodiments≦100 pM to about 1 pM, as measured by assays such as radioligand bindingassays or similar assays known to those skilled in the art.

The term “isolated polynucleotide” as used herein shall mean apolynucleotide of genomic, cDNA, or synthetic origin or some combinationthereof, which by virtue of its origin the “isolated polynucleotide” (1)is not associated with all or a portion of a polynucleotide in which the“isolated polynucleotide” is found in nature, (2) is operably linked toa polynucleotide that it is not linked to in nature, or (3) does notoccur in nature as part of a larger sequence. Polynucleotides inaccordance with the disclosure include the nucleic acid moleculesencoding the heavy chain immunoglobulin molecules shown herein, andnucleic acid molecules encoding the light chain immunoglobulin moleculesshown herein.

The term “isolated protein” referred to herein means a protein of cDNA,recombinant RNA, or synthetic origin or some combination thereof, whichby virtue of its origin, or source of derivation, the “isolated protein”(1) is not associated with proteins found in nature, (2) is free ofother proteins from the same source, e.g., free of murine proteins, (3)is expressed by a cell from a different species, or (4) does not occurin nature.

The term “polypeptide” is used herein as a generic term to refer tonative protein, fragments, or analogs of a polypeptide sequence. Hence,native protein fragments, and analogs are species of the polypeptidegenus. Polypeptides in accordance with the disclosure comprise the heavychain immunoglobulin molecules shown herein, and the light chainimmunoglobulin molecules shown herein, as well as antibody moleculesformed by combinations comprising the heavy chain immunoglobulinmolecules with light chain immunoglobulin molecules, such as kappa lightchain immunoglobulin molecules, and vice versa, as well as fragments andanalogs thereof.

The term “naturally-occurring” as used herein as applied to an objectrefers to the fact that an object can be found in nature. For example, apolypeptide or polynucleotide sequence that is present in an organism(including viruses) that can be isolated from a source in nature andthat has not been intentionally modified by man in the laboratory orotherwise is naturally-occurring.

The term “operably linked” as used herein refers to positions ofcomponents so described are in a relationship permitting them tofunction in their intended manner. A control sequence “operably linked”to a coding sequence is ligated in such a way that expression of thecoding sequence is achieved under conditions compatible with the controlsequences.

The term “control sequence” as used herein refers to polynucleotidesequences that are necessary to effect the expression and processing ofcoding sequences to which they are ligated. The nature of such controlsequences differs depending upon the host organism in prokaryotes, suchcontrol sequences generally include promoter, ribosomal binding site,and transcription termination sequence in eukaryotes, generally, suchcontrol sequences include promoters and transcription terminationsequence. The term “control sequences” is intended to include, at aminimum, all components whose presence is essential for expression andprocessing, and can also include additional components whose presence isadvantageous, for example, leader sequences and fusion partnersequences. The term “polynucleotide” as referred to herein meansnucleotides of at least 10 bases in length, either ribonucleotides ordeoxynucleotides or a modified form of either type of nucleotide. Theterm includes single and double stranded forms of DNA.

The term oligonucleotide referred to herein includes naturallyoccurring, and modified nucleotides linked together by naturallyoccurring, and non-naturally occurring oligonucleotide linkages.Oligonucleotides are a polynucleotide subset generally comprising alength of 200 bases or fewer. In some embodiments, oligonucleotides are10 to 60 bases in length, for example in some embodiments, 12, 13, 14,15, 16, 17, 18, 19, or 20 to 40 bases in length. Oligonucleotides areusually single stranded, e.g., for probes, although oligonucleotides maybe double stranded, e.g., for use in the construction of a gene mutant.Oligonucleotides of the disclosure are either sense or antisenseoligonucleotides.

The term “naturally occurring nucleotides” referred to herein includesdeoxyribonucleotides and ribonucleotides. The term “modifiednucleotides” referred to herein includes nucleotides with modified orsubstituted sugar groups and the like. The term “oligonucleotidelinkages” referred to herein includes oligonucleotide linkages such asphosphorothioate, phosphorodithioate, phosphoroselerloate,phosphorodiselenoate, phosphoroanilothioate, phoshoraniladate,phosphoronmidate, and the like. See e.g., LaPlanche et al. Nucl. AcidsRes. 14:9081 (1986); Stec et al. J. Am. Chem. Soc. 106:6077 (1984),Stein et al. Nucl. Acids Res. 16:3209 (1988), Zon et al. Anti CancerDrug Design 6:539 (1991); Zon et al. Oligonucleotides and Analogues: APractical Approach, pp. 87-108 (F. Eckstein, Ed., Oxford UniversityPress, Oxford England (1991)); Stec et al. U.S. Pat. No. 5,151,510;Uhlmann and Peyman Chemical Reviews 90:543 (1990). An oligonucleotidecan include a label for detection, if desired.

As used herein, the twenty conventional amino acids and theirabbreviations follow conventional usage. See Immunology—A Synthesis (2ndEdition, E. S. Golub and D. R. Gren, Eds., Sinauer Associates,Sunderland7 Mass. (1991)). Stereoisomers (e.g., D-amino acids) of thetwenty conventional amino acids, unnatural amino acids such as α-,α-disubstituted amino acids, N-alkyl amino acids, lactic acid, and otherunconventional amino acids may also be suitable components forpolypeptides of the present disclosure. Examples of unconventional aminoacids include: 4 hydroxyproline, γ-carboxyglutamate,δ-N,N,N-trimethyllysine, δ-N-acetyllysine, O-phosphoserine,N-acetylserine, N-formylmethionine, 3-methylhistidine, 5-hydroxylysine,σ-N-methylarginine, and other similar amino acids and imino acids (e.g.,4-hydroxyproline). In the polypeptide notation used herein, theleft-hand direction is the amino terminal direction and the right-handdirection is the carboxy-terminal direction, in accordance with standardusage and convention.

Similarly, unless specified otherwise, the left-hand end ofsingle-stranded polynucleotide sequences is the 5′ end the left-handdirection of double-stranded polynucleotide sequences is referred to asthe 5′ direction. The direction of 5′ to 3′ addition of nascent RNAtranscripts is referred to as the transcription direction sequenceregions on the DNA strand having the same sequence as the RNA and thatare 5′ to the 5′ end of the RNA transcript are referred to as “upstreamsequences”, sequence regions on the DNA strand having the same sequenceas the RNA and that are 3′ to the 3′ end of the RNA transcript arereferred to as “downstream sequences”.

As applied to polypeptides, the term “substantial identity” means thattwo peptide sequences, when optimally aligned, such as by the programsGAP or BESTFIT using default gap weights, share at least 80 percentsequence identity, for example in some embodiments, at least 90 percentsequence identity, in some embodiments at least 95 percent sequenceidentity, and in some embodiments at least 99 percent sequence identity.

In some embodiments, residue positions that are not identical differ byconservative amino acid substitutions.

As discussed herein, minor variations in the amino acid sequences ofantibodies or immunoglobulin molecules are contemplated as beingencompassed by the present disclosure, providing that the variations inthe amino acid sequence maintain at least 75%, for example in someembodiments at least 80%, 90%, 95%, and in some embodiments 99%. Inparticular, conservative amino acid replacements are contemplated.Conservative replacements are those that take place within a family ofamino acids that are related in their side chains. Genetically encodedamino acids are generally divided into families: (1) acidic amino acidsare aspartate, glutamate; (2) basic amino acids are lysine, arginine,histidine; (3) non-polar amino acids are alanine, valine, leucine,isoleucine, proline, phenylalanine, methionine, tryptophan, and (4)uncharged polar amino acids are glycine, asparagine, glutamine,cysteine, serine, threonine, tyrosine. The hydrophilic amino acidsinclude arginine, asparagine, aspartate, glutamine, glutamate,histidine, lysine, serine, and threonine. The hydrophobic amino acidsinclude alanine, cysteine, isoleucine, leucine, methionine,phenylalanine, proline, tryptophan, tyrosine and valine. Other familiesof amino acids include (i) serine and threonine, which are thealiphatic-hydroxy family; (ii) asparagine and glutamine, which are theamide containing family; (iii) alanine, valine, leucine and isoleucine,which are the aliphatic family; and (iv) phenylalanine, tryptophan, andtyrosine, which are the aromatic family. For example, it is reasonableto expect that an isolated replacement of a leucine with an isoleucineor valine, an aspartate with a glutamate, a threonine with a serine, ora similar replacement of an amino acid with a structurally related aminoacid will not have a major effect on the binding or properties of theresulting molecule, especially if the replacement does not involve anamino acid within a framework site. Whether an amino acid change resultsin a functional peptide can readily be determined by assaying thespecific activity of the polypeptide derivative. Assays are described indetail herein. Fragments or analogs of antibodies or immunoglobulinmolecules can be readily prepared by those of ordinary skill in the art.In some embodiments, amino- and carboxy-termini of fragments or analogsoccur near boundaries of functional domains. Structural and functionaldomains can be identified by comparison of the nucleotide and/or aminoacid sequence data to public or proprietary sequence databases.Computerized comparison methods are used to identify sequence motifs orpredicted protein conformation domains that occur in other proteins ofknown structure and/or function. Methods to identify protein sequencesthat fold into a known three-dimensional structure are known. Bowie etal. Science 253:164 (1991). Thus, the foregoing examples demonstratethat those of skill in the art can recognize sequence motifs andstructural conformations that may be used to define structural andfunctional domains in accordance with the disclosure.

In some embodiments, amino acid substitutions are those that: (1) reducesusceptibility to proteolysis, (2) reduce susceptibility to oxidation,(3) alter binding affinity for forming protein complexes, (4) alterbinding affinities, and (4) confer or modify other physicochemical orfunctional properties of such analogs. Analogs can include variousmuteins of a sequence other than the naturally-occurring peptidesequence. For example, single or multiple amino acid substitutions (forexample, conservative amino acid substitutions) may be made in thenaturally-occurring sequence (for example, in the portion of thepolypeptide outside the domain(s) forming intermolecular contacts. Aconservative amino acid substitution should not substantially change thestructural characteristics of the parent sequence (e.g., a replacementamino acid should not tend to break a helix that occurs in the parentsequence, or disrupt other types of secondary structure thatcharacterizes the parent sequence). Examples of art-recognizedpolypeptide secondary and tertiary structures are described in Proteins,Structures and Molecular Principles (Creighton, Ed., W. H. Freeman andCompany, New York (1984)); Introduction to Protein Structure (C. Brandenand J. Tooze, eds., Garland Publishing, New York, N.Y. (1991)); andThornton et at. Nature 354:105 (1991).

The term “polypeptide fragment” as used herein refers to a polypeptidethat has an amino terminal and/or carboxy-terminal deletion and/or oneor more internal deletion(s), but where the remaining amino acidsequence is identical to the corresponding positions in thenaturally-occurring sequence deduced, for example, from a full lengthcDNA sequence. Fragments typically are at least 5, 6, 8 or 10 aminoacids long, for example in some embodiments at least 14 amino acidslong, in some embodiments at least 20 amino acids long, usually at least50 amino acids long, and in some embodiments at least 70 amino acidslong. The term “analog” as used herein refers to polypeptides that arecomprised of a segment of at least 25 amino acids that has substantialidentity to a portion of a deduced amino acid sequence and that hasspecific binding to EGFR, under suitable binding conditions. Typically,polypeptide analogs comprise a conservative amino acid substitution (oraddition or deletion) with respect to the naturally-occurring sequence.Analogs typically are at least 20 amino acids long, for example in someembodiments at least 50 amino acids long or longer, and can often be aslong as a full-length naturally-occurring polypeptide.

The term “agent” is used herein to denote a chemical compound, a mixtureof chemical compounds, a biological macromolecule, or an extract madefrom biological materials.

As used herein, the terms “label” or “labeled” refers to incorporationof a detectable marker, e.g., by incorporation of a radiolabeled aminoacid or attachment to a polypeptide of biotinyl moieties that can bedetected by marked avidin (e.g., streptavidin containing a fluorescentmarker or enzymatic activity that can be detected by optical orcalorimetric methods). In certain situations, the label or marker canalso be therapeutic. Various methods of labeling polypeptides andglycoproteins are known in the art and may be used. Examples of labelsfor polypeptides include, but are not limited to, the following:radioisotopes or radionuclides (e.g., ³H, ¹⁴C, ¹⁵N, ³⁵S, ⁹⁰Y, ⁹⁹Tc,¹¹¹In, ¹²⁵I, ¹³¹I), fluorescent labels (e.g., a fluorophore, rhodamine,lanthanide phosphors), enzymatic labels (e.g., horseradish peroxidase,p-galactosidase, luciferase, alkaline phosphatase), chemiluminescent,biotinyl groups, predetermined polypeptide epitopes recognized by asecondary reporter (e.g., leucine zipper pair sequences, binding sitesfor secondary antibodies, metal binding domains, epitope tags). In someembodiments, labels are attached by spacer arms of various lengths toreduce potential steric hindrance. The term “pharmaceutical agent ordrug” as used herein refers to a chemical compound or compositioncapable of inducing a desired therapeutic effect when properlyadministered to a patient.

As used herein, “substantially pure” means an object species is thepredominant species present (i.e., on a molar basis it is more abundantthan any other individual species in the composition), and asubstantially purified fraction is a composition wherein the objectspecies comprises at least about 50 percent (on a molar basis) of allmacromolecular species present.

Generally, a substantially pure composition will comprise more thanabout 80 percent of all macromolecular species present in thecomposition, for example, in some embodiments, more than about 85%, 90%,95%, and 99%. In some embodiments, the object species is purified toessential homogeneity (contaminant species cannot be detected in thecomposition by conventional detection methods) wherein the compositionconsists essentially of a single macromolecular species.

The term patient includes human and veterinary subjects.

Other chemistry terms herein are used according to conventional usage inthe art, as exemplified by The McGraw-Hill Dictionary of Chemical Terms(Parker, S., Ed., McGraw-Hill, San Francisco (1985)).

Multispecific Antibodies and Multispecific Activatable Antibodies

Exemplary multispecific antibodies and/or multispecific activatableantibodies of the disclosure include, for example, those shown in theExamples provided herein, and variants thereof.

In some non-limiting embodiments, at least one of the AB in themultispecific antibody is specific for CD3ε and at least one other AB isa binding partner for any target listed in Table 1.

TABLE 1 Exemplary Targets 1-92-LFA-3 CD52 DL44 HVEM LAG-3 STEAP1 Alpha-4CD56 DLK1 Hyaluronidase LIF-R STEAP2 integrin Alpha-V CD64 DLL4 ICOSLewis X TAG-72 integrin alpha4beta1 CD70 DPP-4 IFNalpha LIGHT TAPA1integrin alpha4beta7 CD71 DSG1 IFNbeta LRP4 TGFbeta integrin AGR2 CD74EGFR IFNgamma LRRC26 TIGIT Anti-Lewis-Y EGFRviii IgE MCSP TIM-3 Apelin JCD80 Endothelin B IgE Receptor Mesothelin TLR2 receptor receptor (FceRI)(ETBR) APRIL CD81 ENPP3 IGF MRP4 TLR4 B7-H4 CD86 EpCAM IGF1R MUC1 TLR6BAFF CD95 EPHA2 IL1B Mucin-16 TLR7 (MUC16, CA-125) BTLA CD117 EPHB2 IL1RNa/K ATPase TLR8 C5 CD125 ERBB3 IL2 Neutrophil TLR9 complement elastaseC-242 CD132 F protein of IL11 NGF TMEM31 (IL-2RG) RSV CA9 CD133 FAP IL12Nicastrin TNFalpha CA19-9 CD137 FGF-2 IL12p40 Notch TNFR (Lewis a)Receptors Carbonic CD138 FGF8 IL-12R, Notch 1 TNFRS12A anhydrase 9IL-12Rbeta1 CD2 CD166 FGFR1 IL13 Notch 2 TRAIL-R1 CD3 CD172A FGFR2 IL13RNotch 3 TRAIL-R2 CD6 CD248 FGFR3 IL15 Notch 4 Transferrin CD9 CDH6 FGFR4IL17 NOV Transferrin receptor CD11a CEACAM5 Folate IL18 OSM-R TRK-A(CEA) receptor CD19 CEACAM6 GAL3ST1 IL21 OX-40 TRK-B (NCA-90) CD20CLAUDIN-3 G-CSF IL23 PAR2 uPAR CD22 CLAUDIN-4 G-CSFR IL23R PDGF-AA VAP1CD24 cMet GD2 IL27/IL27R PDGF-BB VCAM-1 (wsx1) CD25 Collagen GITR IL29PDGFRalpha VEGF CD27 Cripto GLUT1 IL-31R PDGFRbeta VEGF-A CD28 CSFRGLUT4 IL31/IL31R PD-1 VEGF-B CD30 CSFR-1 GM-CSF IL2R PD-L1 VEGF-C CD33CTLA-4 GM-CSFR IL4 PD-L2 VEGF-D CD38 CTGF GP IIb/IIIa IL4R Phosphatidyl-VEGFR1 receptors serine CD40 CXCL10 Gp130 IL6, IL6R P1GF VEGFR2 CD40CXCL13 GPIIB/IIIA Insulin PSCA VEGFR3 Receptor CD41 CXCR1 GPNMB JaggedPSMA VISTA Ligands CD44 CXCR2 GRP78 Jagged 1 RAAG12 WISP-1 CD44v6HER2/neu Jagged 2 RAGE WISP-2 CD47 CXCR4 HGF SLC44A4 WISP-3 CD51 CYR61hGH Sphingosine 1 Phosphate

In some non-limiting embodiments, at least one of the AB of themultispecific antibody is or is derived from a sequence set forth in theExamples provided herein.

In some non-limiting embodiments, at least one of the AB of themultispecific antibody is or is derived from an antibody listed in Table2.

TABLE 2 Exemplary sources for ABs Antibody Trade Name (antibody name)Target Avastin ™ (bevacizumab) VEGF Lucentis ™ (ranibizumab) VEGFErbitux ™ (cetuximab) EGFR Vectibix ™ (panitumumab) EGFR Remicade ™(infliximab) TNFα Humira ™ (adalimumab) TNFα Tysabri ™ (natalizumab)Integrinα4 Simulect ™ (basiliximab) IL2R Soliris ™ (eculizumab)Complement C5 Raptiva ™ (efalizumab) CD11a Bexxar ™ (tositumomab) CD20Zevalin ™ (ibritumomab tiuxetan) CD20 Rituxan ™ (rituximab) CD20Ocrelizumab CD20 Arzerra ™ (ofatumumab) CD20 Obinutuzumab CD20 Zenapax ™(daclizumab) CD25 Adcetris ™ (brentuximab vedotin) CD30 Myelotarg ™(gemtuzumab) CD33 Mylotarg ™ (gemtuzumab ozogamicin) CD33 Campath ™(alemtuzumab) CD52 ReoPro ™ (abiciximab) Glycoprotein receptor IIb/IIIaXolair ™ (omalizumab) IgE Herceptin ™ (trastuzumab) Her2 Kadcyla ™(trastuzumab emtansine) Her2 Synagis ™ (palivizumab) F protein of RSVYervoy ™ (ipilimumab) CTLA-4 (tremelimumab) CTLA-4 Hu5c8 CD40L(pertuzumab) Her2-neu (ertumaxomab) CD3/Her2-neu Orencia ™ (abatacept)CTLA-4 (tanezumab) NGF (bavituximab) Phosphatidylserine (zalutumumab)EGFR (mapatumumab) EGFR (matuzumab) EGFR (nimotuzumab) EGFR ICR62 EGFRmAb 528 EGFR CH806 EGFR MDX-447 EGFR/CD64 (edrecolomab) EpCAM RAV12RAAG12 huJ591 PSMA Enbrel ™ (etanercept) TNF-R Amevive ™ (alefacept)1-92-LFA-3 Antril ™, Kineret ™ (ankinra) IL-1Ra GC1008 TGFbeta Notch,e.g., Notch 1 Jagged 1 or Jagged 2 (adecatumumab) EpCAM (figitumumab)IGF1R (tocilizumab) IL-6 receptor Stelara ™ (ustekinumab) IL-12/IL-23Prolia ™ (denosumab) RANKL Keytruda (Pembrolizumab) PD-1 Opdivo(Nivolumab) PD-1 Atezolizumab (MPDL3280A) PD-L1

Exemplary antibodies, activatable antibodies and antigen-bindingfragments thereof that bind CD3ε of the disclosure include the CD3εbinding sequences described in the Examples, for example in Examples 2and 4. Exemplary multispecific antibodies and multispecific activatableantibodies include the multispecific antibodies and multispecificactivatable antibodies described in the Examples, for example in Example4.

In some embodiments, the multispecific activatable antibody includes aheavy chain sequence selected from the group consisting of SEQ ID NOs:446, 452, 454, 456, 460, 462, 464, 466, 470, 472, 476, 478, 480, 486,488, 490, 492, 494, 496, 498, 500, 502, 504, 510, 512, 514, 518, 524,526, 530, 532, 534, 536, 538, 540, 542, 544, and 546.

In some embodiments, the multispecific activatable antibody includes alight chain sequence selected from the group consisting of SEQ ID NOs:448, 450, 458, 468, 474, 482, 484, 508, 516, and 520.

In some embodiments, the multispecific activatable antibody includes aheavy chain sequence selected from the group consisting of SEQ ID NOs:446, 452, 454, 456, 460, 462, 464, 466, 470, 472, 476, 478, 480, 486,488, 490, 492, 494, 496, 498, 500, 502, 504, 510, 512, 514, 518, 524,526, 530, 532, 534, 536, 538, 540, 542, 544, and 546; and a light chainsequence selected from the group consisting of SEQ ID NOs: 448, 450,458, 468, 474, 482, 484, 508, 516, and 520.

In some embodiments, the multispecific activatable antibody includes aheavy chain sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%,97%, 98% or 99% identical to an amino acid sequence selected from thegroup consisting of SEQ ID NOs: 446, 452, 454, 456, 460, 462, 464, 466,470, 472, 476, 478, 480, 486, 488, 490, 492, 494, 496, 498, 500, 502,504, 510, 512, 514, 518, 524, 526, 530, 532, 534, 536, 538, 540, 542,544, and 546.

In some embodiments, the multispecific activatable antibody includes alight chain sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%,97%, 98% or 99% identical to an amino acid sequence selected from thegroup consisting of SEQ ID NOs: 448, 450, 458, 468, 474, 482, 484, 508,516, and 520.

In some embodiments, the multispecific activatable antibody includes aheavy chain sequence that is at least 90%, 91%, 92%, 93%, 94%, 95%, 96%,97%, 98% or 99% identical to an amino acid sequence selected from thegroup consisting of SEQ ID NOs: 446, 452, 454, 456, 460, 462, 464, 466,470, 472, 476, 478, 480, 486, 488, 490, 492, 494, 496, 498, 500, 502,504, 510, 512, 514, 518, 524, 526, 530, 532, 534, 536, 538, 540, 542,544, and 546; and a light chain sequence that is at least 90%, 91%, 92%,93%, 94%, 95%, 96%, 97%, 98% or 99% identical to an amino acid sequenceselected from the group consisting of SEQ ID NOs: 448, 450, 458, 468,474, 482, 484, 508, 516, and 520.

In some embodiments, the multispecific activatable antibody includes theamino acid sequence of SEQ ID NO: 506. In some embodiments, themultispecific activatable antibody includes an amino acid sequence thatis at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% or 99% identicalto the amino acid sequence of SEQ ID NO: 506.

Exemplary ABs that bind CD3ε of the disclosure include the CD3ε bindingsequences shown below:

CD3HvLv (SEQ ID NO: 587)EVQLVESGGGLVQPGGSLKLSCAASGFTFNKYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKNTAYLQMNNLKTEDTAVYYCVRHGNFGNSYISYWAYWGQGTLVTVSSGGGGSGGGGSGGGGSQTVVTQEPSLTVSPGGTVTLTCGSSTGAVTSGYYPNWVQQKPGQAPRGLIGGTKFLAPGT PARFSGSLLGGKAALTLSGVQPEDEAEYYCALWYSNRWVFGGGTKLTVL CD3HvLv(SEQ ID NO: 588) QTVVTQEPSLTVSPGGTVTLTCRSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGTPARFSGSLLGGKAALTLSGVQPEDEAEYYCALWYSNLWVFGGGTKLTVLGGGGSGGGGSGGGGSEVQLVESGGGLVQPGGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDD SKNTAYLQMNNLKTEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS

In some embodiments, the activatable anti-CD3ε antibody includes a heavychain that comprises or is derived from a heavy chain amino acidsequence shown in Table 17. In some embodiments, the activatableanti-CD3ε antibody includes a light chain that comprises or is derivedfrom a light chain amino acid sequence shown in Table 17. In someembodiments, the activatable anti-CD3ε antibody includes a heavy chainthat comprises or is derived from a heavy chain amino acid sequenceshown in Table 17, and a light chain that comprises or is derived from alight chain amino acid sequence shown in Table 17. In some embodiments,the activatable anti-CD3ε antibody includes a combination of a heavychain variable region sequence and a light chain variable regionsequence from the combinations shown in Group A in Table 17. In someembodiments, the activatable anti-CD3ε antibody includes a combinationof a heavy chain variable region sequence and a light chain variableregion sequence from the combinations shown in Group B in Table 17. Insome embodiments, the activatable anti-CD3ε antibody includes acombination of a heavy chain variable region sequence and a light chainvariable region sequence from the combinations shown in Group C in Table17. In some embodiments, the activatable anti-CD3ε antibody includes acombination of a heavy chain variable region sequence and a light chainvariable region sequence from the combinations shown in Group D in Table17. In some embodiments, the activatable anti-CD3ε antibody includes acombination of a heavy chain variable region sequence and a light chainvariable region sequence from the combinations shown in Group E in Table17. In some embodiments, the activatable anti-CD3ε antibody includes acombination of a heavy chain variable region sequence and a light chainvariable region sequence from the combinations shown in Group F in Table17. In some embodiments, the activatable anti-CD3ε antibody includes acombination of a heavy chain variable region sequence and a light chainvariable region sequence from the combinations shown in Group G in Table17. In some embodiments, the activatable anti-CD3ε antibody includes acombination of a heavy chain variable region sequence and a light chainvariable region sequence from the combinations shown in Group H in Table17. In some embodiments, the activatable anti-CD3ε antibody includes acombination of a heavy chain variable region sequence and a light chainvariable region sequence from the combinations shown in Group I in Table17. In some embodiments, the activatable anti-CD3ε antibody includes acombination of a heavy chain variable region sequence and a light chainvariable region sequence from the combinations shown in Group J in Table17. In some embodiments, the activatable anti-CD3ε antibody includes acombination of a heavy chain variable region sequence and a light chainvariable region sequence from the combinations shown in Group K in Table17. In some embodiments, the activatable anti-CD3ε antibody includes acombination of a heavy chain variable region sequence and a light chainvariable region sequence from the combinations shown in Group L in Table17. In some embodiments, the activatable anti-CD3ε antibody includes acombination of a heavy chain variable region sequence and a light chainvariable region sequence from the combinations shown in Group M in Table17. In some embodiments, the activatable anti-CD3ε antibody includes acombination of a heavy chain variable region sequence and a light chainvariable region sequence from the combinations shown in Group N in Table17. In some embodiments, the activatable anti-CD3ε antibody includes acombination of a heavy chain variable region sequence and a light chainvariable region sequence from the combinations shown in Group 0 in Table17. In some embodiments, the activatable anti-CD3ε antibody includes acombination of a heavy chain variable region sequence and a light chainvariable region sequence from the combinations shown in Group P in Table17. In some embodiments, the activatable anti-CD3ε antibody includes acombination of a heavy chain variable region sequence and a light chainvariable region sequence from the combinations shown in Group Q in Table17.

TABLE 17 Variable Heavy Chain Region (VH) and Variable Light Chain Region (VL) Sequences for Activatable Antibodies that Bind CDR Group A VHQIVLTQSPAIMSASPGEKVTMTCSASSSVSYMNWYQQKSGTSPKRWIYDTSKLASGVPAHFRGSGSGTSYSLTISGMEAEDAATYYCQQWSSNPFT (SEQ ID NO: 589) VHQVQLQQSGAELARPGASVKMSCKASGYTFTRYTMHWVKQRPGQGLEWIGYINPSRGYTNYNQKFKDKATLTTDKSSSTAYMQLSSLTSEDSAVYYCARYYDDHYCLDYWGQG (SEQ ID NO: 590) VHQVQLVQSGGGVVQPGRSLRLSCKASGYTFTRYTMHWVRQAPGKGLEWIGYINPSRGYTNYNQKFKDRFTISTDKSKSTAFLQMDSLRPEDTAVYYCARYYDDHYCLDYWGQGTPVTVSS (SEQ ID NO: 591) VHQVQLVESGGGVVQPGRSLRLSCSSSGYTFTRYTMHWVRQAPGKGLEWVAYINPSRGYTNYNQKFKDRFTISRDNSKNTLFLQMDSLRPEDTGVYFCARYYDDHYCLDYWGQGTPVTVSS (SEQ ID NO: 592) VLDIQMTQSPSSLSASVGDRVTITCSASSSVSYMNWYQQTPGKAPKRWIYDTSKLASGVPSRFSGSGSGTDYTFTISSLQPEDIATYYCQQWSSNPFT (SEQ ID NO: 593) VLQIVMTQSPSSLSASVGDRVTITCSASSSVSYMNWYQQTPGKAPKRWIYDTSKLASGVPSRFSGSGSGTDYTFVLDIQMTQSPSSLSASVGDRVTITCSASSSVSYMNWYQQTPGKAPKLLIYDTSKLASGVPSRFSGSGSGTDYTFTISSLQPEDIATYYCQQWSSNPFTFGQGTKLQITR (SEQ ID NO: 595) Group B VHQVQLQQSGAELARPGASVKMSCKASGYTFTRYTMHWVKQRPGQGLEWIGYINPSRGYTNYNQKFKDKATLTTDKSSSTAYMQLSSLTSEDSAVYYCARYYDDHYCLDYWGQGTTLTVSS (SEQ ID NO: 596) VHEVQLVESGGGLVQPGKSLKLSCEASGFTFSGYGMHWVRQAPGRGLESVAYITSSSINIKYADAVKGRFTVSRDNAKNLLFLQMNILKSEDTAMYYCARFDWDKNYWGQGTMVTVSS (SEQ ID NO: 597) VLQIVLTQSPAIMSASPGEKVTMTCRASSSVSYMNWYQQKSGTSPKRWIYDTSKVASGVPYRFSGSGSGTSYSLTISSMEAEDAATYYCQQWSSNPLTFGSGTKLEINR (SEQ ID NO: 598) VLQIVLTQSPAIMSASPGEKVTMTCSASSSVSYMNWYQQKSGTSPKRWIYDTSKLASGVPAHFRGSGSGTSYSLTISGMEAEDAATYYCQQWSSNPFTFGSGTKLEINR (SEQ ID NO: 599) VLDIQMTQSPSSLPASLGDRVTINCQASQDISNYLNWYQQKPGKAPKLLIYYTNKLADGVPSRFSGSGSGRDSSFTISSLESEDIGSYYCQQYYNYPWTFGPGTKLEIKR (SEQ ID NO: 600) Group C VHMGWSWIFLFLLSGTAGVHSQVQLVQSGAEVKKPGASVKVSCKASGYTFISYTMHWVRQAPGQGLEWMGYINPRSGYTHYNQKLKDKATLTADKSASTAYMELSSLRSEDTAVYYCARSAYYDYDGFAYWGQGTLVTVSS(SEQ ID NO: 601) VLMETDTLLLWVLLLWVPGSTGDIQMTQSPSSLSASVGDRVTITCSASSSVSYMNWYQQKPGKAPKRLIYDTSKLASGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQWSSNPPTFGGGTKVEIK (SEQ ID NO: 602)Group D VHEVQLVESGGGLVQPGGSLRLSCAASGYSFTGYTMNWVRQAPGKGLEWVALINPYKGVSTYNQKFKDRFTISVDKSKNTAYLQMNSLRAEDTAVYYCARSGYYGDSDWYFDVWGQGTLVTVSS (SEQ ID NO: 603) VLDIQMTQSPSSLSASVGDRVTITCRASQDIRNYLNWYQQKPGKAPKLLIYYTSRLESGVPSRFSGSGSGTDYTLTISSLQPEDFATYYCQQGNTLPWTFGQGTKVEIK (SEQ ID NO: 604)MGWSCIILFLVATATGVHSEVQLVESGGGLVQPGGSLRLSCATSGYTFTEYTMHWMRQAPGKGLEWVAGINPHCKNGGTSHNQRFMDRFTISVDKSTSTAYMQMNSLRAEDTAVYYCARWRGLNYGFDVRYFDVWGQGTLVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVTSSNFGTQTYTCNVDHKPSNTKVDKTVERKCCVECPPCPAPPVAGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVQFNWYVDGMEVHNAKTKPREEQFNSTFRVVSVLTVVHQDWLNGKEYKCKVSNKGLPAPIEKTISKTKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPMLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 605) HCEVQLVESGGGLVQPGGSLRLSCATSGYTFTEYTMHWMRQAPGKGLEWVAGINPKNGGTSYADSVKGRFTISVDKSKNTLYLQMNSLRAEDTAVYYCARWRGLNYGFDVRYFDVWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHT (SEQ ID NO: 606) HCEVQLVESGGGLVQPGGSLRLSCATSGYTFTEYTMHWMRQAPGKGLEWVAGINPKNGGTSHNQRFMDRFTISVDKSKNTLYLQMNSLRAEDTAVYYCARWRGLNYGFDVRYFDVWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHT (SEQ ID NO: 607) HCEVQLVESGGGLVQPGGSLRLSCATSGYTFTEYTMHWMRQAPGKGLEWVAGINPKNGGTSHNQRFMDRFTLAVDKSKNTLYLQMNSLRAEDTAVYYCARWRGLNYGFDVRYFDVWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHT (SEQ ID NO: 608) HCEVQLVESGGGLVQPGGSLRLSCATSGYTFTEYTMHWMRQAPGKGLEWVAGINPKNGGTSHNQRFMDRFTISVDKSTSTAYMQMNSLRAEDTAVYYCARWRGLNYGFDVRYFDVWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHT (SEQ ID NO: 609) HCEVQLVESGGGLVQPGGSLRLSCATSGYTFTEYTMHWMRQAPGKGLEWVAGINPKNGGTSHNQRFMDRFTISVDKSTSTAYMQMNSLRAEDTAVYYCARWRGLNYGFDVRYFDVWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPP (SEQ ID NO: 610) HCEVQLVESGGGLVQPGGSLRLSCATSGYTFTEYTMHWMRQAPGKGLEWVAGINPKNGGTSHNQRFMDRFTISVDKSKNTLYMELRSLRAEDTAVYYCARWRGLNYGFDVRYFDVWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHT (SEQ ID NO: 611) HCEVQLVESGGGLVQPGGSLRLSCATSGYTFTEYTMHWMRQAPGKGLEWVAGINPKNGGTSHNQRFMDRFTISVDKSKNTLYLQMNSLTSEDSGIYYCARWRGLNYGFDVRYFDVWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHT (SEQ ID NO: 612) HCEVQLVESGGGLVQPGGSLRLSCATSGYTFTEYTMHWMRQAPGKGLEWVAGINPKNGGTSHNQRFMDRFTISVDKSKNTLYLQMNSLRAEDTAVYYCARWRGLNYGFDVRYFDVWGAGTTVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHT (SEQ ID NO: 613) HCEVQLQQSGPELVQPGGSLRLSCATSGYTFTEYTMHWMRQAPGKGLEWVAGINPKNGGTSHNQRFMDRFTISVDKSKNTLYLQMNSLRAEDTAVYYCARWRGLNYGFDVRYFDVWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHT (SEQ ID NO: 614) HCEVQLVESGGGLVKPGASLRLSCATSGYTFTEYTMHWMRQAPGKGLEWVAGINPKNGGTSHNQRFMDRFTISVDKSKNTLYLQMNSLRAEDTAVYYCARWRGLNYGFDVRYFDVWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHT (SEQ ID NO: 615) HCEVQLVESGGGLVQPGGSLKISCKTSGYTFTEYTMHWMRQAPGKGLEWVAGINPKNGGTSHNQRFMDRFTISVDKSKNTLYLQMNSLRAEDTAVYYCARWRGLNYGFDVRYFDVWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHT (SEQ ID NO: 616) HCEVQLVESGGGLVQPGGSLRLSCATSGYTFTEYTMHWMKQSHGKSLEWVAGINPKNGGTSHNQRFMDRFTISVDKSKNTLYLQMNSLRAEDTAVYYCARWRGLNYGFDVRYFDVWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHT (SEQ ID NO: 617) HCEVQLVESGGGLVQPGGSLRLSCATSGYTFTEYTMHWMRQAPGKGLEWIGGFNPKNGGTSHNQRFMDRFTISVDKSKNTLYLQMNSLRAEDTAVYYCARWRGLNYGFDVRYFDVWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHT (SEQ ID NO: 618) LCMGWSCIILFLVATATGVHSDIQMTQSPSSLSASVGDRVTITCRASQDINNYLNWYQQKPGKAPKLLIYYTSTLHSGVPSRFSGSGSGTDYTLTISSLQPEDFATYYCQQGNTLPPTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 619) LCDIQMTQSPSSLSASVGDRVTITCRASQDINNYLNWYQQKPGKAPKLLIYYTSTLESGVPSRFSGSGSGTDYTLTISSLQPEDFATYYCQQGNTLPPTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC(SEQ ID NO: 620) LCDIQMTQSPSSLSASVGDRVTITCRASQDINNYLNWYQQKPGKAPKLLIYYTSTLHSGVPSRFSGSGSGTDYTLTISSLQPEDFATYYCQQGNTLPPTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC(SEQ ID NO: 621) Group E HCMERHWIFLLLLSVTAGVHSQVQLQQSGAELARPGASVKMSCKASGYTFTRYTMHWVKQRPGQGLEWIGYINPSRGYTNYNQKFKDKATLTTDKSSSTAYMQLSSLTSEDSAVYYCARYYDDHYCLDYWGQGTTLTVSSAKTTAPSVYPLAPVCGDTTGSSVTLGCLVKGYFPEPVTLTWNSGSLSSGVHTFPAVLQSDLYTLSSSVTVTSSTWPSQSITCNVAHPASSTKVDKKIEPRGPTIKPCPPCKCPAPNLLGGPSVFIFPPKIKDVLMISLSPIVTCVVVDVSEDDPDVQISWFVNNVEVHTAQTQTHREDYNSTLRVVSALPIQHQDWMSGKEFKCKVNNKDLPAPIERTISKPKGSVRAPQVYVLPPPEEEMTKKQVTLTCMVTDFMPEDIYVEWTNNGKTELNYKNTEPVLDSDGSYFMYSKLRVEKKNWVERNSYSCSVVHEGLHNHHTTKSFSRTPGK (SEQ ID NO: 622) LCMDFQVQIFSFLLISASVIISRGQIVLTQSPAIMSASPGEKVTMTCSASSSVSYMNWYQQKSGTSPKRWIYDTSKLASGVPAHFRGSGSGTSYSLTISGMEAEDAATYYCQQWSSNPFTFGSGTKLEINRADTAPTVSIFPPSSEQLTSGGASVVCFLNNFYPKDINVKWKIDGSERQNGVLNSWTDQDSKDSTYSMSSTLTLTKDEYERHNSYTCEATHKTSTSPIVKSFNRNEC (SEQ ID NO: 623) Group F VHEVQLLESGGGLVQPGGSLRLSCAASGFTFSSFPMAWVRQAPGKGLEWVSTISTSGGRTYYRDSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKFRQYSGGFDYWGQGTLVTVSS (SEQ ID NO: 624) VLDFMLTQPHSVSESPGKTVIISCTLSSGNIENNYVHWYQQRPGRAPTTVIFDDDKRPDGVPDRFSGSIDRSSNSASLTISGLQTEDEADYYCHSYVSSFNVFGGGTKLTVLGQPKAAPSVTLFPPSSEELQ(SEQ ID NO: 625) Group G HCEVQLLESGGGLVQPGGSLRLSCAASGFTFSSFPMAWVRQAPGKGLEWVSTISTSGGRTYYRDSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKFRQYSGGFDYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYASTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 626) VLQAVVTQANSVSTSLGSTVKLSCTLSSGNIENNYVHWYQLYEGRSPTTMIYDDDKRPDGVPDRFSGSIDRSSNSAFLTIHNVAIEDEAIYFCHSYVSSFNVFGGGTKLTVLR (SEQ ID NO: 627) LCQAVVTQANSVSTSLGSTVKLSCTLSSGNIENNYVHWYQLYEGRSPTTMIYDDDKRPDGVPDRFSGSIDRSSNSAFLTIHNVAIEDEAIYFCHSYVSSFNVFGGGTKLTVLRQPKAAPSVTLFPPSSEELQANKATLVCLISDFYPGAVTVAWKADSSPVKAGVETTTPSKQSNNKYAASSYLSLTPEQWKSHRSYSCQVTHEGSTVEKTVAPTECS(SEQ ID NO: 628) Group H VHEVKLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKSSLYLQMNNLKTEDTAMYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 629)VHEVKLVESGGGLVKPGRSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKSILYLQMNNLKTEDTAMYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 630)VHEVKLVESGGGLVKPGRSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKSILYLQMNSLKTEDTAMYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 631)VHEVKLVESGGGLVKPGRSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKSILYLQMNSLKTEDTAMYYCVRHGNFGNSYVSWFAYWGQGTMVTVSS (SEQ ID NO: 632)VLQAVVTOEPSFSVSPGGTVTLTCRSSTGAVTTSNYANWVQQTPGQAFRGLIGGTNKRAPGVPARFSGSLIGDKAALTITGAQADDESIYFCALWYSNLWVFGGGTKLTVL (SEQ ID NO: 633) VLQAVVTOEPSFSVSPGGTVTLTCRSSTGAVTTSNYANWVQQTPGQAFRGLIGGTNKRAPGVPARFSGSILGNKAALTITGAQADDESIYFCALWYSNLWVFGGGTKLTVL (SEQ ID NO: 634) VLQAVVTOEPSFSVSPGGTVTLTCRSSTGAVTTSNYANWVQQTPGQAFRGLIGGTNKRAPGVPARFSGSILGNKAALTITGAQADDESDYYCALWYSNLWVFGGGTKLTVL (SEQ ID NO: 635) Group I VHEVQLVESGGGLVQPGRSLRLSCAASGFTFDDYTMHWVRQAPGKGLEWVSGISWNSGSIGYADSVKGRFTISRDNAKKSLYLQMNSLRAEDTALYYCAKDNSGYGHYYYGMDVWGQGTTVTVAS (SEQ ID NO: 636) VHEVQLVESGGGLVQPGGSLRLSCAATGFTFDDFTMHWVRQAPGKGLEWVSGISWNSGSIGYVDSVKGRFTISRDNAKNSLYLQMNSLRAEDTALYYCAKDNSGYGYYYYGMDVWGQGTTVTVSS (SEQ ID NO: 637) VHQVQLVESGGGVVQPGRSLRLSCAASGFTFRSYAMHWVRQAPGKGLEWVAMVYYDGNNQYYADSVRGRFTISRDNSKNTLYLQMNSLRADDTAVYFCARGPGYNWLDPWGQGTLVTVSS (SEQ ID NO: 638) VHQVQLVESGGGVVQPGRSLRLACVASGFTFRSYGMHWVRQAPGKGLQWVAMIYYDGKNKYYADSVRGRFTISRDNSKNTLYLQMNNLRVEDTAMYFCARGPGYNWLDPWGQGTLVTVSS (SEQ ID NO: 639) VHDVQLVQSGAEVKKPGASVKVSCKASGYTFTRYTMHWVRQAPGQGLEWIGYINPSRGYTNYADSVKGRFTITTDKSTSTAYMELSSLRSEDTATYYCARYYDDHYCLDYWGQGTTVTVSS (SEQ ID NO: 640) VLAEIVMTQSPATLSVSPGERATLSCRASQSVSSNLAWYQQKPGQAPRLLIYGASTRATGIPARFSGSGSGTEFTLTISSLQSEDFAVYYCQHYINWPLTFGGGTKVEIK (SEQ ID NO: 641) VLEIVMTQSPATLSVSPGERATLSCRASHSVSRNSAWYQQKPGQAPRLLIYGASTRATGIPARFSGSGSGTEFTLTISSLQSEDFAIYYCQQYNNWPLTFGGGTKVEIK (SEQ ID NO: 642) VLEIVMTQSPATLSVSPGERATLSCRASQSVSSNLAWYQQKPGQAPRLLIYGASTRATGIPARFSGSGSGTEFTLTISSLQSEDFAVYYCQHYINWPLTFGGGTKVEIK (SEQ ID NO: 643) VLEIVMTQSPATLSVSPGERATLSCRASQSVSRNLAWYQQKPGQAPRLLIYGASTRATGIPARFSGSGSGTDFTLTISSLQSEDFAVYYCQQYNNWPLTFGGGTKVVIK (SEQ ID NO: 644) VLEIVMTQSPATLSVSPGERATLSCRASQRISSNLAWYQQKPGQAPRLLIYGASTRATGSPARFSGSGSGTDFTLTISSLQSEDVAVYYCQQHHNWPLTFGGGTKVEIK (SEQ ID NO: 645) VLDIVLTQSPATLSLSPGERATLSCRASQSVSYMNWYQQKPGKAPKRWIYDTSKVASGVPARFSGSGSGTDYSLTINSLEAEDAATYYCQQWSSNPLTFGGGTKVEIK (SEQ ID NO: 646) Group J VHQVQLVQSGAEVKKPGASVKVSCKASGYTFTRYTMHWVRQAPGQGLEWMGYINPSRGYTNYNQKFKDRVTMTTDTSISTAYMELSRLRSDDTAVYYCARYYDDHYCLDYWGQGTLVTVSS (SEQ ID NO: 647) VHQVQLVQSGAEVKKPGASVKVSCKASGYTFTRYTMHWVRQAPGQGLEWMGYINPSRGYTNYNQKFKDRVTMTTDTSISTAYMELSRLRSDDTAVYYCARYYDDHYSLDYWGQGTLVTVSS (SEQ ID NO: 648) VLEIVLTQSPATLSLSPGERATLSCSASSSVSYMNWYQQKPGQAPRLLIYDTSKLASGVPAHFRGSGSGTDYTLTISSLEPEDFAVYYCQQWSSNPFTFGQGTKVEIK (SEQ ID NO: 649) Group K VHQVQLQQSGAELARPGASVKMSCKASGYTFTRYTMHWVKQRPGQGLEWIGYINPSRGYTNYNQKFKDKATLTTDKSSSTAYMQLSSLTSEDSAVYYCARYYDDHYCLDYWGQGTTLTVSS (SEQ ID NO: 596) VLQIVLTQSPAIMSASPGEKVTMTCSASSSVSYMNWYQQKSGTSPKRWIYDTSKLASGVPAHFRGSGSGTSYSLTISGMEAEDAATYYCQQWSSNPFTFGSGTKLEIN (SEQ ID NO: 650) Group L VHQVQLVQSGSELKKPGASVKMSCKASGYTFTRYTMHWVRQAPGKGLEWIGYINPSRGYTNYNQKFKDRATLTTDKSTSTAYMQLSSLRSEDTAVYYCARYYDDHYSLDYWGQGTLVTVSS (SEQ ID NO: 651) VLQIVLTQSPATLSLSPGERATMSCSASSSVSYMNWYQQKPGKAPKRWIYDTSKLASGVPSRFRGSGSGTDYTLTISSLQPEDFATYYCQQWSSNPFTFGGGTKVEIK(SEQ ID NO: 652) Group M VHEVQLVESGGGLVQPGGSLRLSCAASGFTFSTYAMNWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKDRFTISRDDSKNSLYLQMNSLKTEDTAVYYCARHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 653)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKDRFTISRDDSKNSLYLQMNSLKTEDTAVYYCARHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 654)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFSTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKNSLYLQMNSLKTEDTAVYYCARHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 655)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFSTYAMNWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKDRFTISRDDSKNSLYLQMNSLKTEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 656)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKNSLYLQMNSLKTEDTAVYYCARHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 657)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKDRFTISRDDSKNSLYLQMNSLKTEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 658)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRNKYNNYATEYADSVKDRFTISRDDSKNSLYLQMNSLKTEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 659)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFSTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKNSLYLQMNSLKTEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 660)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKNSLYLQMNSLKTEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 661)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRNKYNNYATEYADSVKDRFTISRDDSKNSLYLQMNSLKTEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 662)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVARTRSKANSYTTYYAASVKGRFTISRDDSKNSLYLQMNSLKTEDTAVYYCARHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 663)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRTRSKANSYTTYYAASVKGRFTISRDDSKNSLYLQMNSLKTEDTAVYYCARHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 664)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRSKYNNYATEYAASVKDRFTISRDDSKNSLYLQMNSLKTEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 665)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRNKYNNYATEYAASVKDRFTISRDDSKNSLYLQMNSLKTEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 666)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRTRSKANSYTTYYAASVKGRFTISRDDSKNSLYLQMNSLKTEDTAVYYCARHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 667)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVARTRSKANSYTTYYAASVKGRFTISRDDSKNSLYLQMNSLKTEDTAVYYCARHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 668)VHQVQLVQSGAEVKKPGASVKVSCKASGYTFTRSTMHWVRQAPGQGLEWIGYINPSSAYTNYNQKFKDRVTITADKSTSTAYMELSSLRSEDTAVYYCASPQVHYDYNGFPYWGQGTLVTVSS (SEQ ID NO: 669) VLQAVVTQEPSLTVSPGGTVTLTCRSSTGAVTTSNYANWFQQKPGQAPRTLIGGTNKRAPWTPARFSGSLLGGKAALTITGAQAEDEADYYCALWYSNLWVFGGGTKLTVLG (SEQ ID NO: 670) VLQAVVTQEPSLTVSPGGTVTLTCRSSTGAVTTSNYANWVQQKPGQAPRTLIGGTNKRAPWTPARFSGSLLGGKAALTITGAQAEDEADYYCALWYSNLWVFGGGTKLTVLG (SEQ ID NO: 671) VLQAVVTQEPSLTVSPGGTVTLTCRSSTGAVTTSNYANWFQEKPGQAPRTLIGGTNKRAPWTPARFSGSLLGGKAALTITGAQAEDEADYYCALWYSNLWVFGGGTKLTVLG (SEQ ID NO: 672) VLQAVVTQEPSLTVSPGGTVTLTCRSSTGAVTTSNYANWFQQKPGQAPRGLIGGTNKRAPWTPARFSGSLLGGKAALTITGAQAEDEADYYCALWYSNLWVFGGGTKLTVLG (SEQ ID NO: 673) VLQAVVTQEPSLTVSPGGTVTLTCRSSTGAVTTSNYANWVQEKPGQAPRTLIGGTNKRAPWTPARFSGSLLGGKAALTITGAQAEDEADYYCALWYSNLWVFGGGTKLTVLG (SEQ ID NO: 674) VLQAVVTQEPSLTVSPGGTVTLTCRSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPWTPARFSGSLLGGKAALTITGAQAEDEADYYCALWYSNLWVFGGGTKLTVLG (SEQ ID NO: 675) VLQAVVTQEPSLTVSPGGTVTLTCRSSTGAVTTSNYANWFQEKPGQAPRGLIGGTNKRAPWTPARFSGSLLGGKAALTITGAQAEDEADYYCALWYSNLWVFGGGTKLTVLG (SEQ ID NO: 676) VLQAVVTQEPSLTVSPGGTVTLTCRSSTGAVTTSNYANWVQEKPGQAPRGLIGGTNKRAPWTPARFSGSLLGGKAALTITGAQAEDEADYYCALWYSNLWVFGGGTKLTVLG (SEQ ID NO: 677) VLQAVVTQEPSLTVSPGGTVTLTCRSSTGAVTTSNYANWVQEKPGQAFRGLIGGTNKRAPWTPARFSGSLLGGKAALTITGAQAEDEADYYCALWYSNLWVFGGGTKLTVLG (SEQ ID NO: 678) VLDIQMTQSPSSLSASVGDRVTITCSASSSVSYMNWYQQKPGKAPKRLIYDSSKLASGVPSRFSGSGSGTEFTLTISSLQPEDFATYYCQQWSRNPPTFGGGTKVEIK (SEQ ID NO: 679) VLDVVMTQSPAIMSAFPGEKVTITCSASSSVSYMNWYQQKPGKAPKRWIYDSSKLASGVPSRFSGSGSGTEFTLTISSLQPEDFATYYCQQWSRNPPTFGGGTKVEIK (SEQ ID NO: 680) Group N VHQVQLVQSGAEVKKPGASVKVSCKASGYTFTRSTMHWVKQAPGQGLEWIGYINPSSAYTNYNQKFKDKATLTADKSSSTAYMQLSSLRSEDTAVYYCARPQVHYDYNGFPYWGQGTLVTVSS (SEQ ID NO: 681) VHQVQLVQSGGGVVQPGRSLRLSCKASGYTFTRSTMHWVRQAPGKGLEWIGYINPSSAYTNYNQKFKDKATLTADKSKNTAYMELSSLRSEDTAVYYCARPQVHYDYNGFPYWGQGTLVTVSS (SEQ ID NO: 682) VHQVQLVQSGGGVVQPGRSLRLSCKASGYTFTRSTMHWVRQAPGKGLEWIGYINPSSAYTNYNQKFKDRFTISADKSKSTAFLQMDSLRPEDTGVYFCARPQVHYDYNGFPYWGQGTPVTVSS (SEQ ID NO: 683) VHQVQLVESGGGVVQPGRSLRLSCKASGYTFTRSTMHWVRQAPGQGLEWIGYINPSSAYTNYNQKFKDRFTISADKSKSTAFLQMDSLRPEDTGVYFCARPQVHYDYNGFPYWGQGTPVTVSS (SEQ ID NO: 684) VHQVQLVQSGAEVKKPGASVKVSCKASGYTFTRSTMHWVRQAPGQRLEWMGYINPSSAYTNYNQKFKDRVTITRDTSASTAYMELSSLRSEDTAVYYCARPQVHYDYNGFPYWGQGTLVTVSS (SEQ ID NO: 685) VHEVQLVESGGGLVKPGGSLRLSCAASGFTFSRSTMHWVRQAPGKGLEWVSYINPSSAYTNYNQKFKDRFTISRDNAKNSLYLQMNSLRAEDTAVYYCARPQVHYDYNGFPYWGQGTLVTVSS(SEQ ID NO: 686)  VLAQDIQMTQSPSSLSASVGDRVTMTCSASSSVSYMNWYQQKPGKAPKLLIYDSSKLASGVPARFSGSGSGTDYTLTISSLQPEDFATYYCQQWSRNPPTFGGGTKVEIK (SEQ ID NO: 687)  VLEIVLTQSPATLSLSPGERATLSCSASSSVSYMNWYQQKPGQAPRLLIYDSSKLASGIPARFSGSGSGTDFTLTISSLEPEDFAVYYCQQWSRNPPTFGGGTKVEIK (SEQ ID NO: 688) VLAQDIQMTQSPSSLSASVGDRVTMTCSASSSVSYMNWYQQKPGKAPKRWIYDSSKLASGVPARFSGSGSGTDYTLTISSLQPEDFATYYCQQWSRNPPTFGGGTKLQIT (SEQ ID NO: 689) VLAQDIQMTQSPSSLSASVGDRVTITCSASSSVSYMNWYQQTPGKAPKRWIYDSSKLASGVPSRFSGSGSGTDFTLTISSLQPEDIATYYCQQWSRNPPTFGQGTKLQIT (SEQ ID NO: 690) Group O VHQVQLVESGGGVVQPGRSLRLSCAASGFKFSGYGMHWVRQAPGKGLEWVAVIWYDGSKKYYVDSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCARQMGYWHFDLWGRGTLVTVSS (SEQ ID NO: 691) VHQVQLVQSGGGVVQSGRSLRLSCAASGFKFSGYGMHWVRQAPGKGLEWVAVIWYDGSKKYYVDSVKGRFTISRDNSKNTLYLQMNSLRGEDTAVYYCARQMGYWHFDLWGRGTLVTVSS (SEQ ID NO: 692) VHQVQLVESGGGVVQPGRSLRLSCAASGFTFRSYGMHWVRQAPGKGLEWVAIIWYDGSKKNYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCARGTGYNWFDPWGQGTLVTVSS (SEQ ID NO: 693) VHQVQLVQSGGGVVQPGRSLRLSCVASGFTFSSYGMHWVRQAPGKGLEWVAAIWYNGRKQDYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCTRGTGYNWFDPWGQGTLVTVSS (SEQ ID NO: 694) VLEIVLTQSPATLSLSPGERATLSCRASQSVSSYLAWYQQKPGQAPRLLIYDASNRATGIPARFSGSGSGTDFTLTISSLEPEDFAVYYCQQRSNWPPLTFGGGTKVEIK (SEQ ID NO: 695) VLEIVLTQSPRTLSLSPGERATLSCRASQSVSSSYLAWYQQKPGQAPRLLIYGASSRATGIPDRFSGSGSGTDFTLTISRLDPEDFAVYYCQQYGSSPITFGQGTRLEIK (SEQ ID NO: 696) VLDILMTQSPSSLSASVGDRVTITCRASQGISSALAWYQQKPGKAPKLLIYYASSLQSGVPSRFSGSGSGTDYTLTISSLQPEDFATYYCQQYYSTLTFGGGTKVEIK (SEQ ID NO: 697) VLDIVMTQSPSSLSASVGDRVTITCRASQGISSALAWYQQKPGKAPKLLIYDASSLGSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQYYSTLTFGGGTKVEIK (SEQ ID NO: 698)  VLDIQMTQSPFSLSASVGDRVTITCWASQGISSYLAWYQQKPAKAPKLFIYYASSLQSGVPSRFSGSGSGTDYTLTISSLQPEDFATYYCQQYYSTLTFGGGTKVEIK (SEQ ID NO: 699) VLDIEMTQSPFSLSASVGDRVTITCWASQGISSYLAWYQQKPAKAPKLFIYYASSLQSGVPSRFSGSGSGTDYTLTISSLQPEDFATYYCQQYYSTLTFGGGTKVEIK (SEQ ID NO: 700) VLEIVLTQSPATLSLSPGERATLSCRASQSVSSYLAWYQQKPGQAPRLLIYDASNRATGIPARFSGSGSGTDFTLTISSLEPEDFAVYYCQQRSNWPWTFGQGTKVEIK (SEQ ID NO: 701) VLAIQLTQSPSSLSASVGDRVTITCRASQGISSALAWYQQKPGKAPKLLIYDASSLESGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQFNSYPITFGQGTRLEIK (SEQ ID NO: 702) Group P VHQVQLVQSGAEVKKPGASVKVSCKASGYTFISYTMHWVRQAPGQGLEWMGYINPRSGYTHYNQKLKDKATLTADKSASTAYMELSSLRSEDTAVYYCARSAYYDYDGFAYWGQGTLVTVSS (SEQ ID NO: 703) VLDIQMTQSPSSLSASVGDRVTITCSASSSVSYMNWYQQKPGKAPKRLIYDTSKLASGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQWSSNPPTFGGGTKVEIK (SEQ ID NO: 704) Group Q VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 705)VHEVQLVESGGGDVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 706)VHEVQLVESGGGKVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 707)VHEVQLVESGGGNVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 708)VHEVQLVESGGGSVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 709)VHEVQLVESGGGLVQPGGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 710)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFSTYAMNWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 711)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMHWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 712)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMSWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 713)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQASGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 714)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRSKANNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 715)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRSKANSYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 716)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRSKYNNYATAYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 717)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRSKYNNYATYYAASVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 718)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 719)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDTSKNTLYLQMNSLRAEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTV (SEQ ID NO: 720) VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTAYLQMNSLRAEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 721)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRDEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 722)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRKEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 723)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRSEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 724)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCARHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 725)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCTRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 726)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGDSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 727)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGQSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 728)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGNTYVSWFAYWGQGTLVTVSS (SEQ ID NO: 729)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGNSYVSWFDYWGQGTLVTVSS (SEQ ID NO: 730)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGNSYVSWFAYWGQGTTVTVSS (SEQ ID NO: 731)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKSLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 732)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTATYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 733)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFSTYAMNWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGDSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 734)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFSTYAMSWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGDSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 735)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFSTYAMNWVRQAPGKGLEWVGRIRSKANNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGDSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 736)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFSTYAMNWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGDSYVSWFDYWGQGTLVTVSS (SEQ ID NO: 737)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFSTYAMSWVRQAPGKGLEWVGRIRSKANNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGDSYVSWFDYWGQGTLVTVSS (SEQ ID NO: 738)VHEVQLVESGGEVKKPGESLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 739)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRQAPGKGLEWVGRIRSNGGSTYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 740) VHEVQLVESGGGLVQPGGSLRLSCAASGFTFNTYAMNWVRKAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 741)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFSTYAMNWVRKAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 742)VHEVQLVESGGGLVQPGGSLRLSCAASGFTFSTYAMNWVRKAPGKGLEWVGRIRSKYNNYATYYADSVKGRFTISRDDSKNTLYLQMNSLRAEDTAVYYCVRHGNFGDSYVSWFAYWGQGTLVTVSS (SEQ ID NO: 743)VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGVPARFSGSLLGGKAALTLSGAQPEDEAEYYCALWYSNLWVFGGGTKLTVL (SEQ ID NO: 744) VLQAVVTQPPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGVPARFSGSLLGGKAALTLSGAQPEDEAEYYCALWYSNLWVFGGGTKLTVL (SEQ ID NO: 745) VLQAVVTQEPSLSVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGVPARFSGSLLGGKAALTLSGAQPEDEAEYYCALWYSNLWVFGGGTKLTVL (SEQ ID NO: 746) VLQAVVTQEPSLTVSPGATVTLTCGSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGVPARFSGSLLGGKAALTLSGAQPEDEAEYYCALWYSNLWVFGGGTKLTVL (SEQ ID NO: 747) VLQAVVTQEPSLTVSPGQTVTLTCGSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGVPARFSGSLLGGKAALTLSGAQPEDEAEYYCALWYSNLWVFGGGTKLTVL (SEQ ID NO: 748) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTSSNYANWVQQKPGQAPRGLIGGTNKRAPGVPARFSGSLLGGKAALTLSGAQPEDEAEYYCALWYSNLWVFGGGTKLTVL (SEQ ID NO: 749) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTSGHYANWVQQKPGQAPRGLIGGTNKRAPGVPARFSGSLLGGKAALTLSGAQPEDEAEYYCALWYSNLWVFGGGTKLTVL (SEQ ID NO: 750) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGQAPRGLIYDTNKRAPGVPARFSGSLLGGKAALTLSGAQPEDEAEYYCALWYSNLWVFGGGTKLTVL (SEQ ID NO: 751) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNNRAPGVPARFSGSLLGGKAALTLSGAQPEDEAEYYCALWYSNLWVFGGGTKLTVL (SEQ ID NO: 752) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRASGVPARFSGSLLGGKAALTLSGAQPEDEAEYYCALWYSNLWVFGGGTKLTVL (SEQ ID NO: 753) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGQAPRGLIGGTSNKHSWTPARFSGSLLGGKAALTLSGAQPEDEAEYYCALWYSNLWVFGGGTKLTVL (SEQ ID NO: 754) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGVPDRFSGSLLGGKAALTLSGAQPEDEAEYYCALWYSNLWVFGGGTKLTVL (SEQ ID NO: 755) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGVPARFSGSKSGGKAALTLSGAQPEDEAEYYCALWYSNLWVFGGGTKLTVL (SEQ ID NO: 756) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGVPARFSGSSSGGKAALTLSGAQPEDEAEYYCALWYSNLWVFGGGTKLTVL (SEQ ID NO: 757) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGVPARFSGSLLGGKAALTISGAQPEDEAEYYCALWYSNLWVFGGGTKLTVL (SEQ ID NO: 758) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGVPARFSGSLLGGKAALTLSGAQAEDEAEYYCALWYSNLWVFGGGTKLTVL (SEQ ID NO: 759) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGVPARFSGSLLGGKAALTLSGAQSEDEAEYYCALWYSNLWVFGGGTKLTVL (SEQ ID NO: 760) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGVPARFSGSLLGGKAALTLSGAQPEDEADYYCALWYSNLWVFGGGTKLTVL (SEQ ID NO: 761) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGVPARFSGSLLGGKAALTLSGAQPEDEAEYYCLLWYSNLWVFGGGTKLTVL (SEQ ID NO: 762) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGVPARFSGSLLGGKAALTLSGAQPEDEAEYYCALWYSNHWVFGGGTKLTVL (SEQ ID NO: 763) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGVPARFSGSLLGGKAALTLSGAQPEDEAEYYCALWYSNLWVFGGGTKLEIKGS (SEQ ID NO: 764) VLEIVMTQSPATLSLSPGERATLSCRSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGVPARFSGSGSGTDFTLTISSLQPEDFAVYYCALWYSNLWVFGGGTKVEIKGS (SEQ ID NO: 765) VLDIVMTQSPDSLAVSLGERATINCKSSTGAVTTSNYANWVQQKPGQPPKGLIGGTNKRAPGVPDRFSGSGSGTDFTLTISSLQAEDVAVYYCALWYSNLWVFGGGTKVEIKGS (SEQ ID NO: 766) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGVPARFSGSLLGGKAALTDSGAQPEDEADYYCALWYSNHWVFGGGTKLTVL (SEQ ID NO: 767) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQEKPGQAPRGLIGGTNKRAPGVPARFSGSLLGGKAALTLSGAQPEDEAEYYCALWYSNLWVFGGGTKLTVL (SEQ ID NO: 768) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGKAPRGLIGGTNKRAPGVPARFSGSLLGGKAALTLSGAQPEDEAEYYCALWYSNLWVFGGGTKLT (SEQ ID NO: 769) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGTAPRGLIGGTNKRAPGVPARFSGSLLGGKAALTLSGAQPEDEAEYYCALWYSNLWVFGGGTKLTVL (SEQ ID NO: 770) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGSPPRGLIGGTNKRAPGVPARFSGSLLGGKAALTLSGAQPEDEAEYYCALWYSNLWVFGGGTKLTVL (SEQ ID NO: 771) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGQPPRGLIGGTNKRAPGVPARFSGSLLGGKAALTLSGAQPEDEAEYYCALWYSNLWVFGGGTKLTVL (SEQ ID NO: 772) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGQGPRGLIGGTNKRAPGVPARFSGSLLGGKAALTLSGAQPEDEAEYYCALWYSNLWVFGGGTKLTVL (SEQ ID NO: 773) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGQSPRGLIGGTNKRAPGVPARFSGSLLGGKAALTLSGAQPEDEAEYYCALWYSNLWVFGGGTKLTVL (SEQ ID NO: 774) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGQDPRGLIGGTNKRAPGVPARFSGSLLGGKAALTLSGAQPEDEAEYYCALWYSNLWVFGGGTKLTVL (SEQ ID NO: 775) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGVPARFSGSLLGGKAALTISGAQPEDEADYYCALWYSNHWVFGGGTKLTVL (SEQ ID NO: 776) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGVPARFSGSLLGGKAALTISGAQPEDEAEYYCALWYSNHWVFGGGTKLTVL (SEQ ID NO: 777) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGKAPRGLIGGTNKRAPGVPARFSGSLLGGKAALTISGAQPEDEAEYYCALWYSNHWVFGGGTKLTVL (SEQ ID NO: 778) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGQSPRGLIGGTNKRAPGVPARFSGSLLGGKAALTISGAQPEDEAEYYCALWYSNHWVFGGGTKLTVL (SEQ ID NO: 779) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGKSPRGLIGGTNKRAPGVPARFSGSLLGGKAALTISGAQPEDEAEYYCALWYSNHWVFGGGTKLTVL (SEQ ID NO: 780) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGKAPRGLIGGTNKRAPGVPARFSGSLLGGKAALTISGAQPEDEADYYCALWYSNHWVFGGGTKLTVL (SEQ ID NO: 781) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGQSPRGLIGGTNKRAPGVPARFSGSLLGGKAALTISGAQPEDEADYYCALWYSNHWVFGGGTKLTVL (SEQ ID NO: 782) VLQAVVTQEPSLTVSPGGTVTLTCGSSTGAVTTSNYANWVQQKPGKSPRGLIGGTNKRAPGVPARFSGSLLGGKAALTISGAQPEDEADYYCALWYSNHWVFGGGTKLTVL (SEQ ID NO: 783)

In some embodiments, the activatable anti-CD3ε antibody comprises or isderived from a commercially available antibody, such as, for example, ananti-CD3 antibody from Boehringer Mannheim Corp. (Indianapolis, Ind.;Cat. No. 1273 485).

In some embodiments, the activatable anti-CD3ε antibody comprises or isderived from an antibody that is manufactured, secreted or otherwiseproduced by a hybridoma, such as, for example, the hybridoma(s)disclosed in U.S. Pat. Nos. 4,361,549 and/or 4,658,019, and deposited atthe American Type Culture Collection under Accession Number ATCC CRL8001 OKT-3.

In some embodiments, the activatable anti-CD3ε antibody comprises or isderived from an antibody that is manufactured, secreted or otherwiseproduced by a hybridoma, such as, for example, the hybridoma(s)disclosed in PCT Publication No. WO 1995/16037, and deposited at theLeibniz-Institut DSMZ—Deutsche Sammlung von Mikroorganismen andZellkulturen GmbH under Accession Number DSM ACC2152.

In some embodiments, the activatable anti-CD3ε antibody comprises or isderived from an antibody that is manufactured, secreted or otherwiseproduced by a hybridoma, such as, for example, the hybridoma(s)disclosed in PCT Publication No. WO 1991/01752, and deposited at theAmerican Type Culture Collection under Accession Number HB 10166.

In some embodiments, the activatable anti-CD3ε antibody includes a CD3εsequence shown in Table 18, a combination of VL CD3ε sequences selectedfrom the group consisting of those combinations shown in Table 18,and/or a combination of VH CD3ε sequences selected from the groupconsisting of those combinations shown in Table 18. In some embodiments,the activatable anti-CD3ε antibody includes a combination of heavy chainCD3ε sequences selected from the group consisting of the combinationsshown in Group R in Table 18. In some embodiments, the activatableanti-CD3ε antibody includes a combination of light chain CD3ε sequencesselected from the group consisting of the combinations shown in Group Rin Table 18. In some embodiments, the activatable anti-CD3ε antibodyincludes a combination of heavy chain CD3ε sequences selected from thegroup consisting of the combinations shown in Group S in Table 18. Insome embodiments, the activatable anti-CD3ε antibody includes acombination of light chain CD3ε sequences selected from the groupconsisting of the combinations shown in Group S in Table 18. In someembodiments, the activatable anti-CD3ε antibody includes a combinationof heavy chain CD3ε sequences selected from the group consisting of thecombinations shown in Group T in Table 18. In some embodiments, theactivatable anti-CD3ε antibody includes a combination of light chainCD3ε sequences selected from the group consisting of the combinationsshown in Group T in Table 18. In some embodiments, the activatableanti-CD3ε antibody includes a combination of heavy chain CD3ε sequencesselected from the group consisting of the combinations shown in Group Uin Table 18. In some embodiments, the activatable anti-CD3ε antibodyincludes a combination of light chain CD3ε sequences selected from thegroup consisting of the combinations shown in Group U in Table 18. Insome embodiments, the activatable anti-CD3ε antibody includes acombination of heavy chain CD3ε sequences selected from the groupconsisting of the combinations shown in Group V in Table 18. In someembodiments, the activatable anti-CD3ε antibody includes a combinationof light chain CD3ε sequences selected from the group consisting of thecombinations shown in Group V in Table 18. In some embodiments, theactivatable anti-CD3ε antibody includes a combination of heavy chainCD3ε sequences selected from the group consisting of the combinationsshown in Group W in Table 18. In some embodiments, the activatableanti-CD3ε antibody includes a combination of light chain CD3ε sequencesselected from the group consisting of the combinations shown in Group Win Table 18. In some embodiments, the activatable anti-CD3ε antibodyincludes a combination of heavy chain CD3ε sequences selected from thegroup consisting of the combinations shown in Group X in Table 18. Insome embodiments, the activatable anti-CD3ε antibody includes acombination of light chain CD3ε sequences selected from the groupconsisting of the combinations shown in Group X in Table 18.

TABLE 18 Additional CDR Sequences for Antibodies and Activatable Antibodiesth at BindCD38Group R VH VL CDR1 CDR2 CDR3 CDR1 CDR2 CDR3 (SEQ ID NO:) (SEQ ID NO:)(SEQ ID NO:) (SEQ ID NO:) (SEQ ID NO:) (SEQ ID NO:) TLSSGNIENYVH DDDKRPDHSYVSSFNV SFPMA TISTSGGRTYYR FRQYSGGFDY (784) (785) (786) (787)DSVKG (788) (789) Group S VH VL CDR1 CDR2 CDR3 CDR1 CDR2 CDR3(SEQ ID NO:) (SEQ ID NO:) (SEQ ID NO:) (SEQ ID NO:) (SEQ ID NO:)(SEQ ID NO:) TLSSGNIE DDDKRPD (785) HSYVSSFNV SFPMA (787) TISTSGGFRQYSGGFDY NNYVH (786) RTYYRDS (789) (790) VKG (788) Group T VH VL CDR1CDR2 CDR3 CDR1 CDR2 CDR3 (SEQ ID NO:) (SEQ ID NO:) (SEQ ID NO:)(SEQ ID NO:) (SEQ ID NO:) (SEQ ID NO:) GFTFNTYA IRSKYNNYAT (792)VRHGNFGNSYV TGAVTTSNY GTN ALWYSNLWV (791) SWFAY (793) (794) (795) (58)Group U VH VL CDR1 CDR2 CDR3 CDR1 CDR2 CDR3 (SEQ ID NO:) (SEQ ID NO:)(SEQ ID NO:) (SEQ ID NO:) (SEQ ID NO:) (SEQ ID NO:) RYTMHYINPSRGYTNYNQKFK YYDDHYSLDY SASSSVSYMN DTSKLAS QQWSSNPFT (796) D (797)(798) (799) (800) (801) Group V VH VL CDR1 CDR2 CDR3 CDR1 CDR2 CDR3(SEQ ID NO:) (SEQ ID NO:) (SEQ ID NO:) (SEQ ID NO:) (SEQ ID NO:)(SEQ ID NO:) SYGMH VISYDGSNKYYADSVK LSPYCTNGVCW RASQTISNYLN AASTLQSQQFNSYPRT (802) G (803) DAFDI (804) (805) (806) (807) DYAMSFIRSKAYGGTTEYAAS QLWLLQDAFDI RASQGISNYLA AASTLQS QQSYSTPPT (808)VKG (809) (810) (811) (806) (812) SRNWWS DIYHSGSTNYNPSLKS GYTSCRDAFDIRASQGIGNYLA WASIRES QQYYSNPQT (813) (814) (815) (816) (817) (818) GYYWSEINHSGSTNYNPSLKS GRGRFLGWLLG RASQGISNYLN DASNLET QQSYSTPYT (819) (820)GSNWFDP (821) (822) (823) (824) GYYWS EINHSGSTNYNPSLKS GPDRMGHGFDIRASQSISSYLN AASSLQS QQSYSSP (819) (820) (825) (826) (92) (827) SNSAAWNRTYYRSKWYNDYAVSV DRRRIAARQYY RASQSVSSNYLA GASSRAT QKYNSAP (828) KS (829)GMDV (830) (831) (832) (833) SYAMG AVSGSGGSTYYADSVK AKFLGHYYGMDRASQSISSYLN AASSLQS LQHNAYPYT (834) G (835) V (836) (826) (92) (837)NPRMGVS HIFPSDAKAHSASLKS ILGEYYPPAWF KSSQSVLYSSNNKN WASTRES QQYLKIPYT(838) (839) DP (840) YLA (841) (842) (843) Group W VH VL CDR1 CDR2 CDR3CDR1 CDR2 CDR3 (SEQ ID NO:) (SEQ ID NO:) (SEQ ID NO:) (SEQ ID NO:)(SEQ ID NO:) (SEQ ID NO:) IYAMN RIRSKYNNYATYYADS HGNFGNSYVSFGSSTGAVTSGYYPN GTKFLAP ALWYSNRWV (844) VKS (845) FAY (846) (847) (848)(849) KYAMN RIRSKYNNYATYYADS HGNFGNSYISY RSSTGAVTSGYYPN ATDMRPSALWYSNRWV (850) VKD (54) WAY (851) (852) (853) (849) SYAMNRIRSKYNNYATYYADS HGNFGNSYLSF GSSTGAVTSGNYPN GTKFLAP VLWYSNRWV (854)VKG (855) WAY (856) (857) (848) (858) RYAMN RIRSKYNNYATYYADS HGNFGNSYLSY(859) VKG (855) FAY (860) VYAMN RIRSKYNNYATYYADS HGNFGNSYLSW (861)VKK (862) WAY (863) KYAMN RIRSKYNNYATYYADS HGNFGNSYISY (850) VKS (845)YAY (864) GYAMN RIRSKYNNYATYYADS HRNFGNSYLSW (865) VKE (866) FAY (867)VYAMN RIRSKYNNYATYYADS HGNFGNSYISW (861) VKK (862) WAY (868) SYAMNRIRSKYNNYATYYADS HGNFGNSYVSW (854) VKG (855) WAY (869) KYAMNRIRSKYNNYATYYADS HGNFGNSYISY (850) VKD (54) WAY (851) IYAMNRIRSKYNNYATYYADS HGNFGNSYLSF (844) VKS (845) FAY (846) KYAMNRIRSKYNNYATYYADS HGNFGNSYISY (850) VKD (54) WAY (851) SYAMNRIRSKYNNYATYYADS HGNFGNSYLSF (854) VKG (855) WAY (856) GSSTGAVTSGYYPNGTKFLAP ALWYSNRWV (847) (848) (849) RSSTGAVTSGYYPN ATDMRPS ALWYSNRWV(852) (853) (849) GSSTGAVTSGNYPN GTKFLAP VLWYSNRWV (857) (848) (858)Group X VH VL CDR1 CDR2 CDR3 CDR1 CDR2 CDR3 (SEQ ID NO:) (SEQ ID NO:)(SEQ ID NO:) (SEQ ID NO:) (SEQ ID NO:) (SEQ ID NO:) GYGMHVIWYDGSKKYYVDSVK QMGYWHFDL RASQSVSSYLA DASNRAT QQRSNWPPLT (870) G (871)(872) (873) (874) (875) SYGMH IIWYDGSKKNYADSVK GTGYNWFDP RASQSVSSSYLAGASSRAT QQYGSSPIT (802) G (872) (873) (874) (832) (875) SYGMHIIWYDGSKKNYADSVK GTGYNWFDP RASQGISSALA YASSLQS QQYYSTLTF (802) G (872)(873) (876) (877) (888) SYGMH IIWYDGSKKNYADSVK GTGYNWFDP RASQGISSALADASSLGS QQYYSTLTF (802) G (872) (873) (876) (889) (888) SYGMHIIWYDGSKKNYADSVK GTGYNWFDP WASQGISSYLA YASSLQS QQYYSTLTF (802) G (872)(873) (890) (877) (888) SYGMH AIWYNGRKQDYADSVK GTGYNWFDP RASQSVSSYLADASNRAT QQRSNWPWT (802) G (891) (873) (873) (874) (892) SYGMHAIWYNGRKQDYADSVK GTGYNWFDP RASQGISSALA DASSLES QQFNSYPIT (802) G (891)(873) (876) (893) (894)

The ABs in the multispecific activatable antibodies of the disclosurespecifically bind at least a mammalian target. In some embodiments, suchABs bind mammalian CD3ε. In some embodiments, such ABs bind a humantarget. In some embodiments, such ABs bind a non-human primate target.Also included in the disclosure are ABs that bind to the same epitope asan antibody of the disclosure and/or an activated activatable antibodydescribed herein. Also included in the disclosure are ABs that competewith an antibody and/or an activated activatable antibody describedherein for binding to a target, e.g., a human target. Also included inthe disclosure are ABs that cross-compete with an antibody and/or anactivated activatable antibody described herein for binding to a target,e.g., a human target.

In some embodiments, the ABs in the multispecific activatable antibodiesof the disclosure specifically bind at least a CD3ε target, such as, forexample, mammalian CD3ε. In some embodiments, such ABs bind mammalianCD3ε. In some embodiments, such ABs bind human CD3ε. In someembodiments, such ABs bind non-human primate CD3ε. Also included in thedisclosure are ABs that bind to the same CD3ε epitope as an antibody ofthe disclosure and/or an activated activatable antibody describedherein. Also included in the disclosure are ABs that compete with ananti-CD3ε antibody and/or an activated anti-CD3ε activatable antibodydescribed herein for binding to a CD3ε target, e.g., human CD3ε. Alsoincluded in the disclosure are ABs that cross-compete with an anti-CD3εantibody and/or an activated anti-CD3ε activatable antibody describedherein for binding to a CD3ε target, e.g., human CD3ε.

In some embodiments, at least one of the AB of the multispecificantibody and/or multispecific activatable antibody binds EpidermalGrowth Factor Receptor (EGFR). In some embodiments, the AB that bindsEGFR includes one or more of the heavy chain and/or light chainsequences shown below.

C225v5 Antibody Heavy Chain Nucleotide Sequence: (SEQ ID NO: 100)CAGGTGCAGCTGAAACAGAGCGGCCCGGGCCTGGTGCAGCCGAGCCAGAGCCTGAGCATTACCTGCACCGTGAGCGGCTTTAGCCTGACCAACTATGGCGTGCATTGGGTGCGCCAGAGCCCGGGCAAAGGCCTGGAATGGCTGGGCGTGATTTGGAGCGGCGGCAACACCGATTATAACACCCCGTTTACCAGCCGCCTGAGCATTAACAAAGATAACAGCAAAAGCCAGGTGTTTTTTAAAATGAACAGCCTGCAAAGCCAGGATACCGCGATTTATTATTGCGCGCGCGCGCTGACCTATTATGATTATGAATTTGCGTATTGGGGCCAGGGCACCCTGGTGACCGTGAGCGCGGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACAACAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGATGAACTGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA C225v5 Antibody Heavy Chain Amino Acid Sequence (SEQ ID NO: 101)QVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSQDTAIYYCARALTYYDYEFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK* C225v5 Antibody Light Chain Nucleotide Sequence: (SEQ ID NO: 102)CAGATCTTGCTGACCCAGAGCCCGGTGATTCTGAGCGTGAGCCCGGGCGAACGTGTGAGCTTTAGCTGCCGCGCGAGCCAGAGCATTGGCACCAACATTCATTGGTATCAGCAGCGCACCAACGGCAGCCCGCGCCTGCTGATTAAATATGCGAGCGAAAGCATTAGCGGCATTCCGAGCCGCTTTAGCGGCAGCGGCAGCGGCACCGATTTTACCCTGAGCATTAACAGCGTGGAAAGCGAAGATATTGCGGATTATTATTGCCAGCAGAACAACAACTGGCCGACCACCTTTGGCGCGGGCACCAAACTGGAACTGAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG C225v5 Antibody Light Chain Amino Acid Sequence: (SEQ ID NO: 103)QILLTQSPVILSVSPGERVSFSCRASQSIGTNIHWYQQRTNGSPRLLIKYASESISGIPSRFSGSGSGTDFTLSINSVESEDIADYYCQQNNNWPTTFGAGTKLELKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC* C225v4 Antibody Heavy Chain Nucleotide Sequence: (SEQ ID NO: 104)CAGGTGCAGCTGAAACAGAGCGGCCCGGGCCTGGTGCAGCCGAGCCAGAGCCTGAGCATTACCTGCACCGTGAGCGGCTTTAGCCTGACCAACTATGGCGTGCATTGGGTGCGCCAGAGCCCGGGCAAAGGCCTGGAATGGCTGGGCGTGATTTGGAGCGGCGGCAACACCGATTATAACACCCCGTTTACCAGCCGCCTGAGCATTAACAAAGATAACAGCAAAAGCCAGGTGTTTTTTAAAATGAACAGCCTGCAAAGCAACGATACCGCGATTTATTATTGCGCGCGCGCGCTGACCTATTATGATTATGAATTTGCGTATTGGGGCCAGGGCACCCTGGTGACCGTGAGCGCGGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACAACAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGATGAACTGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA C225v4 Antibody Heavy Chain Amino Acid Sequence: (SEQ ID NO: 105)QVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSNDTAIYYCARALTYYDYEFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK* C225v6 Antibody Heavy Chain Nucleotide Sequence: (SEQ ID NO: 106)CAGGTGCAGCTGAAACAGAGCGGCCCGGGCCTGGTGCAGCCGAGCCAGAGCCTGAGCATTACCTGCACCGTGAGCGGCTTTAGCCTGACCAACTATGGCGTGCATTGGGTGCGCCAGAGCCCGGGCAAAGGCCTGGAATGGCTGGGCGTGATTTGGAGCGGCGGCAACACCGATTATAACACCCCGTTTACCAGCCGCCTGAGCATTAACAAAGATAACAGCAAAAGCCAGGTGTTTTTTAAAATGAACAGCCTGCAAAGCCAGGATACCGCGATTTATTATTGCGCGCGCGCGCTGACCTATTATGATTATGAATTTGCGTATTGGGGCCAGGGCACCCTGGTGACCGTGAGCGCGGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACGCCAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGATGAACTGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA]C225v6 Antibody Heavy Chain Amino Acid Sequence (SEQ ID NO: 107)QVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSQDTAIYYCARALTYYDYEFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYASTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK* C225Antibody Heavy Chain Amino Acid Sequence (SEQ ID NO: 523)QVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSQDTAIYYCARALTYYDYEFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK* 

In some embodiments, at least one of the AB of the multispecificantibody and/or multispecific activatable antibody binds interleukin 6receptor (IL-6R). In some embodiments, the AB that binds IL-6R includesone or more of the heavy chain and/or light chain sequences shown below.

Av1 Antibody Heavy Chain Amino Acid Sequence: (SEQ ID NO: 108)QVQLQESGPGLVRPSQTLSLTCTVSGYSITSDHAWSWVRQPPGRGLEWIGYISYSGITTYNPSLKSRVTISRDNSKNTLYLQMNSLRAEDTAVYYCARSLARTTAMDYWGQGSLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK Av1 Antibody Light Chain Amino Acid Sequence: (SEQ ID NO: 109)DIQMTQSPSSLSASVGDRVTITCRASQDISSYLNWYQQKPGKAPKLLIYYTSRLHSGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQGNTLPYTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC 

In some embodiments, the AB binds a Jagged target, e.g., Jagged 1,Jagged 2 or both Jagged 1 and Jagged 2. In some embodiments, the AB thatbinds a Jagged target includes one or more of the heavy chain and/orlight chain sequences shown below.

4D11 Light Chain sequence: (SEQ ID NO: 110)DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQTVVAPPLFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC 4D11 Heavy Chain sequence: (SEQ ID NO: 111)EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSSIDPEGRQTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKDIGGRSAFDYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK 4D11v2 Heavy Chain sequence (SEQ ID NO: 112)EVHLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSSIDPEGRQTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKDIGGRSAFDYWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK 4D11v2 Light Chain Sequence (SEQ ID NO: 113)DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQTVVAPPLFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLXKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC 

In some embodiments, at least one of the AB of the multispecificantibody and/or multispecific activatable antibody a Jagged target andincludes one or more of the variable heavy chain and/or variable lightchain sequences shown below.

Variable Light Chain Amino Sequence Lc4 (SEQ ID NO: 114)DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSVVAPLTFGQGTKVEIKR  Variable Heavy Chain Amino Sequence Hc4(SEQ ID NO: 115)EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSSIEQMGWQTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKDIGGRSAFDYWGQGTLVTVSS Variable Light Chain Amino Sequence Lc5 (SEQ ID NO: 116)DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSVVAPLTFGQGTKVEIKR  Variable Heavy Chain Amino Sequence Hc5(SEQ ID NO: 117)EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSSIEQMGWQTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKSPPYHGQFDYWGQGTLVTVSS Variable Light Chain Amino Sequence Lc7 (SEQ ID NO: 118)DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSVVAPLTFGQGTKVEIKR  Variable Heavy Chain Amino Sequence Hc7(SEQ ID NO: 119)EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSSIEQMGWQTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKSPPFFGQFDYWGQGTLVTVSS Variable Light Chain Amino Sequence Lc8 (SEQ ID NO: 120)DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSVVAPLTFGQGTKVEIKR  Variable Heavy Chain Amino Sequence Hc8(SEQ ID NO: 121)EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSSIEQMGWQTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKHIGRTNPFDYWGQGTLVTVSS Variable Light Chain Amino Sequence Lc13 (SEQ ID NO: 122)DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSVVAPLTFGQGTKVEIKR Variable Heavy Chain Amino Sequence Hc13 (SEQ ID NO: 123)EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSSIEQMGWQTEYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKSAAAFDYWGQGTLVTVSS Variable Light Chain Amino Sequence Lc16 (SEQ ID NO: 124)DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSVVAPLTFGQGTKVEIKR Variable Heavy Chain Amino Sequence Hc16 (SEQ ID NO: 125)EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSSIEQMGWQTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKSPPYYGQFDYWGQGTLVTVSS Variable Light Chain Amino Sequence Lc19 (SEQ ID NO: 126)DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSVVAPLTFGQGTKVEIKR Variable Heavy Chain Amino Sequence Hc19 (SEQ ID NO: 127)EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSSIEQMGWQTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKSPPFFGQFDYWGQGTLVTVSS Variable Light Chain Amino Sequence Lc21 (SEQ ID NO: 128)DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSVVAPLTFGQGTKVEIKR Variable Heavy Chain Amino Sequence Hc21 (SEQ ID NO: 129)EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSSIEQMGWQTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKDIGGRSAFDYWGQGTLVTVSS Variable Light Chain Amino Sequence Lc24 (SEQ ID NO: 130)DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSVVAPLTFGQGTKVEIKR Variable Heavy Chain Amino Sequence Hc24 (SEQ ID NO: 131)EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSSIEEMGWQTLYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKSAAAFDYWGQGTLVTVSS Variable Light Chain Amino Sequence Lc26 (SEQ ID NO: 132)DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSVVAPLTFGQGTKVEIKR Variable Heavy Chain Amino Sequence Hc26 (SEQ ID NO: 133)EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSSIEQMGWQTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKDIGGRSAFDYWGQGTLVTVSS Variable Light Chain Amino Sequence Lc27 (SEQ ID NO: 134)DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSVVAPLTFGQGTKVEIKR Variable Heavy Chain Amino Sequence Hc27 (SEQ ID NO: 135)EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSSIEQMGWQTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKSPPFYGQFDYWGQGTLVTVSS Variable Light Chain Amino Sequence Lc28 (SEQ ID NO: 136)DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSVVAPLTFGQGTKVEIKR Variable Heavy Chain Amino Sequence Hc28 (SEQ ID NO: 137)EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSSIEQMGWQTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKSPPFFGQFDYWGQGTLVTVSS Variable Light Chain Amino Sequence Lc30 (SEQ ID NO: 138)DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSVVAPLTFGQGTKVEIKR Variable Heavy Chain Amino Sequence Hc30 (SEQ ID NO: 139)EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSSIEEMGWQTLYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYAKSAAAFDYWGQGTLVTVSS Variable Light Chain Amino Sequence Lc31 (SEQ ID NO: 140)DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSVVAPLTFGQGTKVEIKR Variable Heavy Chain Amino Sequence Hc31 (SEQ ID NO: 141)EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSSIEQMGWQTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKDIGGRSAFDYWGQGTLVTVSS Variable Light Chain Amino Sequence Lc32 (SEQ ID NO: 142)DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSVVAPLTFGQGTKVEIKR Variable Heavy Chain Amino Sequence Hc32 (SEQ ID NO: 143)EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSSIDPEGWQTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKSAAAFDYWGQGTLVTVSS Variable Light Chain Amino Sequence Lc37 (SEQ ID NO: 144)DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSVVAPLTFGQGTKVEIKR Variable Heavy Chain Amino Sequence Hc37 (SEQ ID NO: 145)EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSSIEQMGWQTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKSPPHNGQFDYWGQGTLVTVSS Variable Light Chain Amino Sequence Lc39 (SEQ ID NO: 146)DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSVVAPLTFGQGTKVEIKR Variable Heavy Chain Amino Sequence Hc39 (SEQ ID NO: 147)EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSSIEQMGWQTEYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKSAAAFDYWGQGTLVTVSS Variable Light Chain Amino Sequence Lc40 (SEQ ID NO: 148)DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSVVAPLTFGQGTKVEIKR  Heavy Chain Amino Sequence Hc40(SEQ ID NO: 149)EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSSIEQMGWQTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKSPPFFGQFDYWGQGTLVTVSS Variable Light Chain Amino Sequence Lc47 (SEQ ID NO: 150)DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSVVAPLTFGQGTKVEIKR Variable Heavy Chain Amino Sequence Hc47 (SEQ ID NO: 151)EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSSIDEMGWQTEYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKSAAAFDYWGQGTLVTVSS  Variable 4B2 Light Chain(SEQ ID NO: 152)DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQTLDAPPQFGQGTKVEIKR  Variable 4B2 Heavy Chain(SEQ ID NO: 153)EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSSIEQMGWQTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKDIGGRSAFDYWGQGTLVTVSS  Variable 4D11 Light Chain(SEQ ID NO: 154)DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQTVVAPPLFGQGTKVEIKR  Variable 4D11 Heavy Chain(SEQ ID NO: 155)EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSSIDPEGRQTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKDIGGRSAFDYWGQGTLVTVSS  Variable 4E7 Light Chain(SEQ ID NO: 156)DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQSLVAPLTFGQGTKVEIKR  Variable 4E7 Heavy Chain(SEQ ID NO: 157)EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSSIEEMGWQTKYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKSAAAFDYWGQGTLVTVSS  Variable 4E11 Light Chain(SEQ ID NO: 158)DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQALDAPLMFGQGTKVEIKR  Variable 4E11 Heavy Chain(SEQ ID NO: 159)EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSSIEPMGQLTEYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKDIGGRSAFDYWGQGTLVTVSS  Variable 6B7 Light Chain(SEQ ID NO: 160)DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQALVAPLTFGQGTKVEIKR  Variable 6B7 Heavy Chain(SEQ ID NO: 161)EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSSIDEMGWQTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKSAAAFDYWGQGTLVTVSS  Variable 6F8 Light Chain(SEQ ID NO: 162)DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKAPKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQALVAPLTFGQGTKVEIKR  Variable 6F8 Heavy Chain(SEQ ID NO: 163)EVQLLESGGGLVQPGGSLRLSCAASGFTFSSYAMSWVRQAPGKGLEWVSSIDEMGWQTYYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCAKSAAAFDYWGQGTLVTVSS 

Antibodies and/or activatable antibodies of the disclosure specificallybind a given target, e.g., a human target protein such as human CD3ε.Also included in the disclosure are anti-CD3ε antibodies, activatableantibodies, multispecific antibodies and/or multispecific activatableantibodies that bind to the same epitope as the anti-CD3ε antibodies,activatable antibodies, multispecific antibodies and/or multispecificactivatable antibodies described herein. Also included in the disclosureare anti-CD3ε antibodies, activatable antibodies, multispecificantibodies and/or multispecific activatable antibodies that compete withan anti-CD3ε antibody, activatable antibody, multispecific antibodyand/or multispecific activatable antibody described herein for bindingto CD3ε, e.g., human CD3ε. Also included in the disclosure are anti-CD3εantibodies, activatable antibodies, multispecific antibodies and/ormultispecific activatable antibodies that cross-compete with ananti-CD3ε antibody, activatable antibody, multispecific antibody and/ormultispecific activatable antibody described herein for binding to CD3ε,e.g., human CD3ε.

Those skilled in the art will recognize that it is possible todetermine, without undue experimentation, if an anti-CD3ε antibody, ananti-CD3ε activatable antibody, a multispecific antibody and/or amultispecific activatable antibody has the same or similar specificityas an anti-CD3ε antibody, an anti-CD3ε activatable antibody, amultispecific antibody and/or multispecific activatable antibody of thedisclosure by ascertaining whether the former prevents the latter frombinding to a target. If the anti-CD3ε antibody, the anti-CD3εactivatable antibody, multispecific antibody and/or a multispecificactivatable antibody being tested competes with the anti-CD3ε antibody,the anti-CD3ε activatable antibody, the multispecific antibody and/or amultispecific activatable antibody of the disclosure, as shown by adecrease in binding by the anti-CD3ε antibody, the anti-CD3ε activatableantibody, the multispecific antibody and/or a multispecific activatableantibody of the disclosure, then the anti-CD3ε antibody, the anti-CD3εactivatable antibody, the two multispecific antibodies and/ormultispecific activatable antibodies bind to the same, or a closelyrelated, epitope.

One embodiment for determining whether an anti-CD3ε antibody, ananti-CD3ε activatable antibody, a multispecific antibody and/or amultispecific activatable antibody has the same or similar specificityas an anti-CD3ε antibody, an anti-CD3ε activatable antibody, amultispecific antibody and/or a multispecific activatable antibody ofthe disclosure is to pre-incubate the anti-CD3ε antibody, the anti-CD3εactivatable antibody, the multispecific antibody and/or a multispecificactivatable antibody of the disclosure with soluble target with which itis normally reactive, and then add the anti-CD3ε antibody, the anti-CD3εactivatable antibody, the multispecific antibody and/or a multispecificactivatable antibody being tested to determine if the anti-CD3εantibody, the anti-CD3ε activatable antibody, the multispecific antibodyand/or a multispecific activatable antibody being tested is inhibited inits ability to bind the target. If the anti-CD3ε antibody, the anti-CD3εactivatable antibody, the multispecific antibody and/or a multispecificactivatable antibody being tested is inhibited then, in all likelihood,it has the same, or functionally equivalent, epitopic specificity as theanti-CD3ε antibody, the anti-CD3ε activatable antibody, themultispecific antibody and/or a multispecific activatable antibody ofthe disclosure.

An anti-CD3ε antibody, an anti-CD3ε activatable antibody, multispecificantibody and/or a multispecific activatable antibody is generated, forexample, using the procedures described in the Examples provided below.An anti-CD3ε antibody, an anti-CD3ε activatable antibody, multispecificantibody and/or a multispecific activatable antibody can also begenerated using any of a number of art-recognized techniques forantibody production and/or purification.

Antibody fragments, such as Fv, F(ab′)₂ and Fab, for use in an anti-CD3εantibody, an anti-CD3ε activatable antibody, a multispecific antibodyand/or a multispecific activatable antibody may be prepared by cleavageof the intact protein, e.g., by protease or chemical cleavage. In someembodiments, a truncated gene is designed. For example, a chimeric geneencoding a portion of the F(ab′)₂ fragment would include DNA sequencesencoding the CH1 domain and hinge region of the H chain, followed by atranslational stop codon to yield the truncated molecule.

Expression vectors include plasmids, retroviruses, YACs, EBV derivedepisomes, and the like. A convenient vector is one that encodes afunctionally complete human CH or CL immunoglobulin sequence, withappropriate restriction sites engineered so that any VH or VL sequencecan be easily inserted and expressed. In such vectors, splicing usuallyoccurs between the splice donor site in the inserted J region and thesplice acceptor site preceding the human C region, and also at thesplice regions that occur within the human CH exons. Polyadenylation andtranscription termination occur at native chromosomal sites downstreamof the coding regions. The resulting antibody may be joined to anystrong promoter, including retroviral LTRs, e.g., SV-40 early promoter,(Okayama et al. Mol. Cell. Bio. 3:280 (1983)), Rous sarcoma virus LTR(Gorman et al. P.N.A.S. 79:6777 (1982)), and moloney murine leukemiavirus LTR (Grosschedl et al. Cell 41:885 (1985)). Also, as will beappreciated, native Ig promoters and the like may be used.

Further, anti-CD3ε antibodies, anti-CD3ε activatable antibodies,multispecific antibodies and/or multispecific activatable antibodies canbe generated through display type technologies, including, withoutlimitation, phage display, retroviral display, ribosomal display, andother techniques, using techniques well known in the art and theresulting molecules can be subjected to additional maturation, such asaffinity maturation, as such techniques are well known in the art.Wright et al. Crit, Reviews in Immunol. 12125-168 (1992), Hanes andPlückthun PNAS USA 94:4937-4942 (1997) (ribosomal display), Parmley andSmith Gene 73:305-318 (1988) (phage display), Scott, TIBS, vol.17:241-245 (1992), Cwirla et al. PNAS USA 87:6378-6382 (1990), Russel etal. Nucl. Acids Research 21:1081-1085 (1993), Hoganboom et al. Immunol.Reviews 130:43-68 (1992), Chiswell and McCafferty TIBTECH; 10:80-8A(1992), and U.S. Pat. No. 5,733,743.

It can be desirable to modify the anti-CD3ε antibody, the anti-CD3εactivatable antibody, the multispecific antibody and/or multispecificactivatable antibody of the disclosure with respect to effectorfunction, so as to enhance or reduce such function to improve theeffectiveness of the antibody in recruiting CD3⁺ cells to one or moretargets that are associated with a disease to be treated, such as, e.g.targets that are expressed on tumor cells. For example, cysteineresidue(s) can be introduced into the Fc region, thereby allowinginterchain disulfide bond formation in this region. The homodimericantibody thus generated can have improved internalization capabilityand/or increased complement-mediated cell killing and antibody-dependentcellular cytotoxicity (ADCC). (See Caron et al., J. Exp Med., 176:1191-1195 (1992) and Shopes, J. Immunol., 148: 2918-2922 (1992)). Insome embodiments, an antibody can be engineered that has dual Fc regionsand can thereby have enhanced complement lysis and ADCC capabilities.(See Stevenson et al., Anti-Cancer Drug Design, 3: 219-230 (1989)). Insome embodiments, Fc mutations are made to remove glycosylation sites,thereby reducing Fc function.

Activatable Antibodies and Multispecific Activatable Antibodies

The activatable antibodies provided herein contain an antibody orantibody fragment thereof (collectively referred to as AB throughout thedisclosure) that specifically binds a target and/or a epitope, whereinthe AB is modified by a masking moiety (MM). The multispecificactivatable antibodies provided herein contain at least a first antibodyor antibody fragment thereof (collectively referred to as AB1 throughoutthe disclosure) that specifically binds a first target and/or a firstepitope and a second antibody or antibody fragment thereof (collectivelyreferred to as AB2 throughout the disclosure) that specifically binds asecond target and/or a second epitope, wherein at least one of the AB ismodified by a masking moiety (MM). In some embodiments, each AB in amultispecific activatable antibody is modified by its own maskingmoiety.

When the AB of an activatable antibody or at least one of the AB in amultispecific activatable antibody is modified with a MM and is in thepresence of its target, specific binding of the AB to its target isreduced or inhibited, as compared to the specific binding of the AB notmodified with an MM or the specific binding of the parental AB to thetarget.

The K_(d) of the AB modified with a MM towards the target is at least 5,10, 20, 25, 40, 50, 100, 250, 500, 1,000, 2,500, 5,000, 10,000, 50,000,100,000, 500,000, 1,000,000, 5,000,000, 10,000,000, 50,000,000 orgreater, or between 5-10, 10-100, 10-1,000, 10-10,000, 10-100,000,10-1,000,000, 10-10,000,000, 100-1,000, 100-10,000, 100-100,000,100-1,000,000, 100-10,000,000, 1,000-10,000, 1,000-100,000,1,000-1,000,000, 1000-10,000,000, 10,000-100,000, 10,000-1,000,000,10,000-10,000,000, 100,000-1,000,000, or 100,000-10,000,000 timesgreater than the K_(d) of the AB not modified with an MM or of theparental AB towards the target. Conversely, the binding affinity of theAB modified with a MM towards the target is at least 5, 10, 20, 25, 40,50, 100, 250, 500, 1,000, 2,500, 5,000, 10,000, 50,000, 100,000,500,000, 1,000,000, 5,000,000, 10,000,000, 50,000,000 or greater, orbetween 5-10, 10-100, 10-1,000, 10-10,000, 10-100,000, 10-1,000,000,10-10,000,000, 100-1,000, 100-10,000, 100-100,000, 100-1,000,000,100-10,000,000, 1,000-10,000, 1,000-100,000, 1,000-1,000,000,1000-10,000,000, 10,000-100,000, 10,000-1,000,000, 10,000-10,000,000,100,000-1,000,000, or 100,000-10,000,000 times lower than the bindingaffinity of the AB not modified with an MM or of the parental AB towardsthe target.

The dissociation constant (K_(d)) of the MM towards the AB in theactivatable antibody is generally greater than the K_(d) of the ABtowards the target. The dissociation constant (K_(d)) of the MM towardsat least one of the AB in the multispecific activatable antibody isgenerally greater than the K_(d) of the AB towards the target. The K_(d)of the MM towards the AB can be at least 5, 10, 25, 50, 100, 250, 500,1,000, 2,500, 5,000, 10,000, 100,000, 1,000,000 or even 10,000,000 timesgreater than the K_(d) of the AB towards the target. Conversely, thebinding affinity of the MM towards the AB is generally lower than thebinding affinity of the AB towards the target. The binding affinity ofMM towards the AB can be at least 5, 10, 25, 50, 100, 250, 500, 1,000,2,500, 5,000, 10,000, 100,000, 1,000,000 or even 10,000,000 times lowerthan the binding affinity of the AB towards the target.

When the AB in the activatable antibody is modified with a MM and is inthe presence of the target, specific binding of the AB to its target isreduced or inhibited, as compared to the specific binding of the AB notmodified with an MM or the specific binding of the parental AB to thetarget. When at least one of the AB in the multispecific activatableantibody is modified with a MM and is in the presence of the target,specific binding of the AB to its target is reduced or inhibited, ascompared to the specific binding of the AB not modified with an MM orthe specific binding of the parental AB to the target. When compared tothe binding of the AB not modified with an MM or the binding of theparental AB to the target, the AB's ability to bind the target whenmodified with an MM can be reduced by at least 50%, 60%, 70%, 80%, 90%,92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% and even 100% for at least 2, 4,6, 8, 12, 28, 24, 30, 36, 48, 60, 72, 84, or 96 hours, or 5, 10, 15, 30,45, 60, 90, 120, 150, or 180 days, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11,or 12 months or more when measured in vivo or in an in vitro assay.

The MM inhibits the binding of the AB in the activatable antibody to itstarget. The MM inhibits the binding of at least one of the AB in themultispecific activatable antibody to its target. The MM binds theantigen binding domain of the AB and inhibits binding of the AB to itstarget. The MM can sterically inhibit the binding of the AB to thetarget. The MM can allosterically inhibit the binding of the AB to itstarget. In these embodiments when the AB is modified or coupled to a MMand in the presence of target, there is no binding or substantially nobinding of the AB to the target, or no more than 0.001%, 0.01%, 0.1%,1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%,or 50% binding of the AB to the target, as compared to the binding ofthe AB not modified with an MM, the parental AB, or the AB not coupledto an MM to the target, for at least 2, 4, 6, 8, 12, 28, 24, 30, 36, 48,60, 72, 84, or 96 hours, or 5, 10, 15, 30, 45, 60, 90, 120, 150, or 180days, or 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months or longer whenmeasured in vivo or in an in vitro assay.

When the AB in an activatable antibody is coupled to or modified by aMM, the MM ‘masks’ or reduces or otherwise inhibits the specific bindingof the AB to its target. When the AB in an activatable antibody iscoupled to or modified by a MM, such coupling or modification can effecta structural change that reduces or inhibits the ability of the AB tospecifically bind its target.

When at least one of the AB in a multispecific activatable antibody iscoupled to or modified by a MM, the MM ‘masks’ or reduces or otherwiseinhibits the specific binding of the AB to its target. When at least oneof the AB in a multispecific activatable antibody is coupled to ormodified by a MM, such coupling or modification can effect a structuralchange that reduces or inhibits the ability of the AB to specificallybind its target.

In an activatable antibody, when the AB is coupled to or modified withan MM, at least a portion of the activatable antibody can be representedby the following formulae (in order from an amino (N) terminal region tocarboxyl (C) terminal region:

-   -   (MM)-(AB)    -   (AB)-(MM)    -   (MM)-L-(AB)    -   (AB)-L-(MM)        where MM is a masking moiety, the AB is an antibody or antibody        fragment thereof, and the L is a linker. In many embodiments, it        may be desirable to insert one or more linkers, e.g., flexible        linkers, into the composition so as to provide for flexibility.

In a multispecific activatable antibody, when at least one AB is coupledto or modified with an MM, at least a portion of the multispecificactivatable antibody can be represented by the following formulae (inorder from an amino (N) terminal region to carboxyl (C) terminal region:

-   -   (MM)-(AB)    -   (AB)-(MM)    -   (MM)-L-(AB)    -   (AB)-L-(MM)        where MM is a masking moiety, the AB is an antibody or antibody        fragment thereof, and the L is a linker. In many embodiments, it        may be desirable to insert one or more linkers, e.g., flexible        linkers, into the composition so as to provide for flexibility.

In certain embodiments, the MM is not a natural binding partner of theAB. In some embodiments, the MM contains no or substantially no homologyto any natural binding partner of the AB. In other embodiments the MM isno more than 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%,65%, 70%, 75%, or 80% similar to any natural binding partner of the AB.In some embodiments, the MM is no more than 5%, 10%, 15%, 20%, 25%, 30%,35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, or 80% identical to anynatural binding partner of the AB. In some embodiments, the MM is nomore than 25% identical to any natural binding partner of the AB. Insome embodiments, the MM is no more than 50% identical to any naturalbinding partner of the AB. In some embodiments, the MM is no more than20% identical to any natural binding partner of the AB. In someembodiments, the MM is no more than 10% identical to any natural bindingpartner of the AB.

In some embodiments, the activatable antibodies and/or the multispecificactivatable antibodies include an AB that is modified by an MM and alsoincludes one or more cleavable moieties (CM). Such activatableantibodies and/or multispecific activatable antibodies exhibitactivatable/switchable binding, to the AB's target. Activatableantibodies and/or multispecific activatable antibodies generally includeat least one antibody or antibody fragment (AB), modified by or coupledto a masking moiety (MM) and a modifiable or cleavable moiety (CM). Insome embodiments, the CM contains an amino acid sequence that serves asa substrate for a protease of interest.

The elements of the activatable antibodies and/or multispecificactivatable antibodies are arranged so that each MM and CM arepositioned such that in a cleaved (or relatively active) state and inthe presence of a target, the corresponding AB binds a target, while inan uncleaved (or relatively inactive) state in the presence of thetarget, specific binding of the AB to its target, is reduced orinhibited. The specific binding of the AB to its target can be reduceddue to the inhibition or masking of the AB's ability to specificallybind its target by the MM.

The K_(d) of each AB modified with a MM and a CM towards the target, isat least 5, 10, 20, 25, 40, 50, 100, 250, 500, 1,000, 2,500, 5,000,10,000, 50,000, 100,000, 500,000, 1,000,000, 5,000,000, 10,000,000,50,000,000 or greater, or between 5-10, 10-100, 10-1,000, 10-10,000,10-100,000, 10-1,000,000, 10-10,000,000, 100-1,000, 100-10,000,100-100,000, 100-1,000,000, 100-10,000,000, 1,000-10,000, 1,000-100,000,1,000-1,000,000, 1000-10,000,000, 10,000-100,000, 10,000-1,000,000,10,000-10,000,000, 100,000-1,000,000, or 100,000-10,000,000 timesgreater than the K_(d) of the AB not modified with an MM and a CM or ofthe parental AB towards the target. Conversely, the binding affinity ofeach AB modified with a MM and a CM towards the target, is at least 5,10, 20, 25, 40, 50, 100, 250, 500, 1,000, 2,500, 5,000, 10,000, 50,000,100,000, 500,000, 1,000,000, 5,000,000, 10,000,000, 50,000,000 orgreater, or between 5-10, 10-100, 10-1,000, 10-10,000, 10-100,000,10-1,000,000, 10-10,000,000, 100-1,000, 100-10,000, 100-100,000,100-1,000,000, 100-10,000,000, 1,000-10,000, 1,000-100,000,1,000-1,000,000, 1000-10,000,000, 10,000-100,000, 10,000-1,000,000,10,000-10,000,000, 100,000-1,000,000, or 100,000-10,000,000 times lowerthan the binding affinity of the AB not modified with an MM and a CM orof the parental AB towards the target.

When at least one AB is modified with a MM and a CM and is in thepresence of the target but not in the presence of a modifying agent (forexample a protease), specific binding of that AB to its target, isreduced or inhibited, as compared to the specific binding of the AB notmodified with an MM and a CM or of the parental AB to the target. Whencompared to the binding of the parental AB or the binding of an AB notmodified with an MM and a CM to its target, the AB's ability to bind thetarget when modified with an MM and a CM can be reduced by at least 50%,60%, 70%, 80%, 90%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% and even 100%for at least 2, 4, 6, 8, 12, 28, 24, 30, 36, 48, 60, 72, 84, or 96 hoursor 5, 10, 15, 30, 45, 60, 90, 120, 150, or 180 days, or 1, 2, 3, 4, 5,6, 7, 8, 9, 10, 11, or 12 months or longer when measured in vivo or inan in vitro assay.

As used herein, the term cleaved state refers to the condition of theactivatable antibodies and/or multispecific activatable antibodiesfollowing modification of the CM by a protease. The term uncleavedstate, as used herein, refers to the condition of the activatableantibodies and/or multispecific activatable antibodies in the absence ofcleavage of the CM by a protease. As discussed above, the term“activatable antibodies” is used herein to refer to an activatableantibody in both its uncleaved (native) state, as well as in its cleavedstate. It will be apparent to the ordinarily skilled artisan that insome embodiments, a cleaved activatable antibody may lack an MM due tocleavage of the CM by protease, resulting in release of at least the MM(e.g., where the MM is not joined to the activatable antibodies by acovalent bond (e.g., a disulfide bond between cysteine residues). Asdiscussed above, the term “multispecific activatable antibodies” is usedherein to refer to a multispecific activatable antibody in both itsuncleaved (native) state, as well as in its cleaved state. It will beapparent to the ordinarily skilled artisan that in some embodiments, acleaved multispecific activatable antibody may lack an MM due tocleavage of the CM by protease, resulting in release of at least the MM(e.g., where the MM is not joined to the multispecific activatableantibodies by a covalent bond (e.g., a disulfide bond between cysteineresidues).

By activatable or switchable is meant that the activatable antibodyand/or multispecific activatable antibody exhibits a first level ofbinding to a target when the activatable antibody is in a inhibited,masked or uncleaved state (i.e., a first conformation), and a secondlevel of binding to the target, when the activatable antibody is in theuninhibited, unmasked and/or cleaved state (i.e., a secondconformation), where the second level of target binding is greater thanthe first level of binding. In general, the access of target to thecorresponding AB of the activatable antibody and/or multispecificactivatable antibody is greater in the presence of a cleaving agentcapable of cleaving the CM than in the absence of such a cleaving agent.Thus, when the activatable antibody and/or multispecific activatableantibody is in the uncleaved state, at least one AB is inhibited fromtarget binding and can be masked from target binding (i.e., the firstconformation is such the AB cannot bind the target), and when theactivatable antibody is in the cleaved state the AB is not inhibited oris unmasked to target binding.

The CM and AB of the activatable antibodies are selected so that thefirst AB represents a binding moiety for CD3ε, and the CM represents asubstrate for a protease that is produced by a tumor that is inproximity to cells that express CD3ε and/or produced by a tumor that isco-localized with CD3ε at a treatment site or diagnostic site in asubject. The activatable antibodies disclosed herein find particular usewhere, for example, a protease capable of cleaving a site in the CM ispresent at relatively higher levels in CD3ε-containing tissue of atreatment site or diagnostic site than in tissue of non-treatment sites(for example in healthy tissue).

The CM and AB of the activatable antibodies and/or multispecificactivatable antibodies are selected so that the first AB represents abinding moiety for a first target and/or epitope, and the CM representsa substrate for a protease that is produced by a tumor that is inproximity to cells that express the target and/or produced by a tumorthat is co-localized with the target at a treatment site or diagnosticsite in a subject. The activatable antibodies and/or multispecificactivatable antibodies disclosed herein find particular use where, forexample, a protease capable of cleaving a site in the CM is present atrelatively higher levels in target-containing tissue of a treatment siteor diagnostic site than in tissue of non-treatment sites (for example inhealthy tissue).

In some embodiments, activatable antibodies provide for reduced toxicityand/or adverse side effects that could otherwise result from binding ofthe AB at non-treatment sites if the AB were not masked or otherwiseinhibited from binding CD3ε.

In general, an activatable antibody can be designed by selecting atfirst AB of interest and constructing the remainder of the activatableantibody so that, when conformationally constrained, the MM provides formasking of the AB or reduction of binding of the AB to CD3ε. Structuraldesign criteria can be to be taken into account to provide for thisfunctional feature.

In some embodiments, multispecific activatable antibodies provide forreduced toxicity and/or adverse side effects that could otherwise resultfrom binding of the first AB at non-treatment sites if the AB were notmasked or otherwise inhibited from binding its target.

In general, a multispecific activatable antibody can be designed byselecting at first AB of interest and constructing the remainder of theactivatable antibody so that, when conformationally constrained, the MMprovides for masking of the AB or reduction of binding of the AB to itstarget. Structural design criteria can be to be taken into account toprovide for this functional feature.

Activatable antibodies and/or multispecific activatable antibodiesexhibiting a switchable phenotype of a desired dynamic range for targetbinding in an inhibited versus an uninhibited conformation are provided.Dynamic range generally refers to a ratio of (a) a maximum detectedlevel of a parameter under a first set of conditions to (b) a minimumdetected value of that parameter under a second set of conditions. Forexample, in the context of an activatable antibody and/or amultispecific activatable antibody, the dynamic range refers to theratio of (a) a maximum detected level of target protein, binding to anactivatable antibody and/or a multispecific activatable antibody in thepresence of protease capable of cleaving the CM of the activatableantibodies to (b) a minimum detected level of target protein, binding toan activatable antibody and/or a multispecific activatable antibody inthe absence of the protease. The dynamic range of an activatableantibody and/or a multispecific activatable antibody can be calculatedas the ratio of the dissociation constant of an activatable antibodyand/or a multispecific activatable antibody cleaving agent (e.g.,enzyme) treatment to the dissociation constant of the activatableantibodies cleaving agent treatment. The greater the dynamic range of anactivatable antibody and/or multispecific activatable antibody, thebetter the switchable phenotype of the activatable antibody. Activatableantibodies having relatively higher dynamic range values (e.g., greaterthan 1) exhibit more desirable switching phenotypes such that targetprotein binding by the activatable antibodies occurs to a greater extent(e.g., predominantly occurs) in the presence of a cleaving agent (e.g.,enzyme) capable of cleaving the CM of the activatable antibodies than inthe absence of a cleaving agent.

Activatable antibodies and/or multispecific activatable antibodies canbe provided in a variety of structural configurations. Exemplaryformulae for at least a portion of an activatable antibody and/or amultispecific activatable antibody are provided below. It isspecifically contemplated that the N- to C-terminal order of the AB, thecorresponding MM and CM may be reversed within an activatable antibody.It is also specifically contemplated that the CM and MM may overlap inamino acid sequence, e.g., such that the CM is contained within the MM.

For example, at least a portion of the activatable antibody activatableantibodies can be represented by the following formula (in order from anamino (N) terminal region to carboxyl (C) terminal region:

-   -   (MM)-(CM)-(AB)    -   (AB)-(CM)-(MM)        where MM is a masking moiety, CM is a cleavable moiety, and AB        is an antibody or fragment thereof. It should be noted that        although MM and CM are indicated as distinct components in the        formulae above, in all exemplary embodiments (including        formulae) disclosed herein it is contemplated that the amino        acid sequences of the MM and the CM could overlap, e.g., such        that the CM is completely or partially contained within the MM.        In addition, the formulae above provide for additional amino        acid sequences that may be positioned N-terminal or C-terminal        to the activatable antibodies elements.

For example, at least a portion of the multispecific activatableantibodies can be represented by the following formula (in order from anamino (N) terminal region to carboxyl (C) terminal region:

-   -   (MM)-(CM)-(AB)    -   (AB)-(CM)-(MM)        where MM is a masking moiety, CM is a cleavable moiety, and AB        is a first antibody or fragment thereof. It should be noted that        although MM and CM are indicated as distinct components in the        formulae above, in all exemplary embodiments (including        formulae) disclosed herein it is contemplated that the amino        acid sequences of the MM and the CM could overlap, e.g., such        that the CM is completely or partially contained within the MM.        In addition, the formulae above provide for additional amino        acid sequences that may be positioned N-terminal or C-terminal        to the activatable antibodies elements.

In certain embodiments, the MM is not a natural binding partner of theAB. In some embodiments, the MM contains no or substantially no homologyto any natural binding partner of the AB. In other embodiments the MM isno more than 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%,65%, 70%, 75%, or 80% similar to any natural binding partner of the AB.In some embodiments, the MM is no more than 5%, 10%, 15%, 20%, 25%, 30%,35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, or 80% identical to anynatural binding partner of the AB. In some embodiments, the MM is nomore than 50% identical to any natural binding partner of the AB. Insome embodiments, the MM is no more than 25% identical to any naturalbinding partner of the AB. In some embodiments, the MM is no more than20% identical to any natural binding partner of the AB. In someembodiments, the MM is no more than 10% identical to any natural bindingpartner of the AB.

In many embodiments it may be desirable to insert one or more linkers,e.g., flexible linkers, into the activatable antibody and/or themultispecific activatable antibody construct so as to provide forflexibility at one or more of the MM-CM junction, the CM-AB junction, orboth. For example, the AB, MM, and/or CM may not contain a sufficientnumber of residues (e.g., Gly, Ser, Asp, Asn, especially Gly and Ser,particularly Gly) to provide the desired flexibility. As such, theswitchable phenotype of such activatable antibody and/or multispecificactivatable antibody constructs may benefit from introduction of one ormore amino acids to provide for a flexible linker. In addition, asdescribed below, where the activatable antibody and/or the multispecificactivatable antibody is provided as a conformationally constrainedconstruct, a flexible linker can be operably inserted to facilitateformation and maintenance of a cyclic structure in the uncleavedactivatable antibody and/or the uncleaved multispecific activatableantibody.

For example, in certain embodiments an activatable antibody and/or amultispecific activatable antibody comprises one of the followingformulae (where the formula below represent an amino acid sequence ineither N- to C-terminal direction or C- to N-terminal direction):

-   -   (MM)-L1-(CM)-(AB)    -   (MM)-(CM)-L2-(AB)    -   (MM)-L1-(CM)-L2-(AB)        wherein MM, CM, and AB are as defined above; wherein L1 and L2        are each independently and optionally present or absent, are the        same or different flexible linkers that include at least 1        flexible amino acid (e.g., Gly). In addition, the formulae above        provide for additional amino acid sequences that may be        positioned N-terminal or C-terminal to the activatable antibody        and/or the multispecific activatable antibodies elements.        Examples include, but are not limited to, targeting moieties        (e.g., a ligand for a receptor of a cell present in a target        tissue) and serum half-life extending moieties (e.g.,        polypeptides that bind serum proteins, such as immunoglobulin        (e.g., IgG) or serum albumin (e.g., human serum albumin (HAS)).

In some non-limiting embodiments, at least one of the AB in themultispecific activatable antibody is a binding partner for any targetlisted in Table 1.

In some non-limiting embodiments, at least one of the AB in themultispecific activatable antibody comprises, is or is derived from asequence set forth in the Examples provided herein.

In some non-limiting embodiments, at least one of the AB in themultispecific activatable antibody comprises, is or is derived from asequence set forth in Example 2 and/or Example 4 in the Examplesprovided herein.

In some non-limiting embodiments, at least one of the AB in themultispecific activatable antibody is or is derived from an antibodylisted in Table 2.

In some embodiments, the masking moiety is selected for use with aspecific antibody or antibody fragment. For example, suitable maskingmoieties for use with antibodies that bind EGFR include MMs that includethe sequence CISPRG (SEQ ID NO: 164). By way of non-limiting examples,the MM can include a sequence such as CISPRGC (SEQ ID NO: 165); CISPRGCG(SEQ ID NO: 166); CISPRGCPDGPYVMY (SEQ ID NO: 167); CISPRGCPDGPYVM (SEQID NO: 168), CISPRGCEPGTYVPT (SEQ ID NO: 169) and CISPRGCPGQIWHPP (SEQID NO: 170). Other suitable masking moieties include any of theEGFR-specific masks disclosed in PCT Publication No. WO 2010/081173,such as, by way of non-limiting example, GSHCLIPINMGAPSC (SEQ ID NO:171); CISPRGCGGSSASQSGQGSHCLIPINMGAPSC (SEQ ID NO: 172);CNHHYFYTCGCISPRGCPG (SEQ ID NO: 173); ADHVFWGSYGCISPRGCPG (SEQ ID NO:174); CHHVYWGHCGCISPRGCPG (SEQ ID NO: 175); CPHFTTTSCGCISPRGCPG (SEQ IDNO: 176); CNHHYHYYCGCISPRGCPG (SEQ ID NO: 177); CPHVSFGSCGCISPRGCPG (SEQID NO: 178); CPYYTLSYCGCISPRGCPG (SEQ ID NO: 179); CNHVYFGTCGCISPRGCPG(SEQ ID NO: 180); CNHFTLTTCGCISPRGCPG (SEQ ID NO: 181);CHHFTLTTCGCISPRGCPG (SEQ ID NO: 182); YNPCATPMCCISPRGCPG (SEQ ID NO:183); CNHHYFYTCGCISPRGCG (SEQ ID NO: 184); CNHHYHYYCGCISPRGCG (SEQ IDNO: 185); CNHVYFGTCGCISPRGCG (SEQ ID NO: 186); CHHVYWGHCGCISPRGCG (SEQID NO: 187); CPHFTTTSCGCISPRGCG (SEQ ID NO: 188); CNHFTLTTCGCISPRGCG(SEQ ID NO: 189); CHHFTLTTCGCISPRGCG (SEQ ID NO: 190);CPYYTLSYCGCISPRGCG (SEQ ID NO: 191); CPHVSFGSCGCISPRGCG (SEQ ID NO:192); ADHVFWGSYGCISPRGCG (SEQ ID NO: 193); YNPCATPMCCISPRGCG (SEQ ID NO:194); CHHVYWGHCGCISPRGCG (SEQ ID NO: 195);C(N/P)H(HN/F)(Y/T)(F/W/T/L)(Y/G/T/S)(T/S/Y/H)CGCISPRGCG (SEQ ID NO:196); CISPRGCGQPIPSVK (SEQ ID NO: 197); CISPRGCTQPYHVSR (SEQ ID NO:198); and/or CISPRGCNAVSGLGS (SEQ ID NO: 199).

Suitable masking moieties for use with antibodies that bind a Jaggedtarget, e.g., Jagged 1 and/or Jagged 2, include, by way of non-limitingexample, masking moieties that include a sequence such asQGQSGQCNIWLVGGDCRGWQG (SEQ ID NO: 200); QGQSGQGQQQWCNIWINGGDCRGWNG (SEQID NO: 201); PWCMQRQDFLRCPQP (SEQ ID NO: 202); QLGLPAYMCTFECLR (SEQ IDNO: 203); CNLWVSGGDCGGLQG (SEQ ID NO: 204); SCSLWTSGSCLPHSP (SEQ ID NO:205); YCLQLPHYMQAMCGR (SEQ ID NO: 206); CFLYSCTDVSYWNNT (SEQ ID NO:207); PWCMQRQDYLRCPQP (SEQ ID NO: 208); CNLWISGGDCRGLAG (SEQ ID NO:209); CNLWVSGGDCRGVQG (SEQ ID NO: 210); CNLWVSGGDCRGLRG (SEQ ID NO:211); CNLWISGGDCRGLPG (SEQ ID NO: 212); CNLWVSGGDCRDAPW (SEQ ID NO:213); CNLWVSGGDCRDLLG (SEQ ID NO: 214); CNLWVSGGDCRGLQG (SEQ ID NO:215); CNLWLHGGDCRGWQG (SEQ ID NO: 216); CNIWLVGGDCRGWQG (SEQ ID NO:217); CTTWFCGGDCGVMRG (SEQ ID NO: 218); CNIWGPSVDCGALLG (SEQ ID NO:219); CNIWVNGGDCRSFEG (SEQ ID NO: 220); YCLNLPRYMQDMCWA (SEQ ID NO:221); YCLALPHYMQADCAR (SEQ ID NO: 222); CFLYSCGDVSYWGSA (SEQ ID NO:223); CYLYSCTDSAFWNNR (SEQ ID NO: 224); CYLYSCNDVSYWSNT (SEQ ID NO:225); CFLYSCTDVSYW (SEQ ID NO: 226); CFLYSCTDVAYWNSA (SEQ ID NO: 227);CFLYSCTDVSYWGDT (SEQ ID NO: 228); CFLYSCTDVSYWGNS (SEQ ID NO: 229);CFLYSCTDVAYWNNT (SEQ ID NO: 230); CFLYSCGDVSYWGNPGLS (SEQ ID NO: 231);CFLYSCTDVAYWSGL (SEQ ID NO: 232); CYLYSCTDGSYWNST (SEQ ID NO: 233);CFLYSCSDVSYWGNI (SEQ ID NO: 234); CFLYSCTDVAYW (SEQ ID NO: 235);CFLYSCTDVSYWGST (SEQ ID NO: 236); CFLYSCTDVAYWGDT (SEQ ID NO: 237);GCNIWLNGGDCRGWVDPLQG (SEQ ID NO: 238); GCNIWLVGGDCRGWIGDTNG (SEQ ID NO:239); GCNIWLVGGDCRGWIEDSNG (SEQ ID NO: 240); GCNIWANGGDCRGWIDNIDG (SEQID NO: 241); GCNIWLVGGDCRGWLGEAVG (SEQ ID NO: 242); GCNIWLVGGDCRGWLEEAVG(SEQ ID NO: 243); GGPALCNIWLNGGDCRGWSG (SEQ ID NO: 244);GAPVFCNIWLNGGDCRGWMG (SEQ ID NO: 245); GQQQWCNIWINGGDCRGWNG (SEQ ID NO:246); GKSEFCNIWLNGGDCRGWIG (SEQ ID NO: 247); GTPGGCNIWANGGDCRGWEG (SEQID NO: 248); GASQYCNLWINGGDCRGWRG (SEQ ID NO: 249); GCNIWLVGGDCRPWVEGG(SEQ ID NO: 250); GCNIWAVGGDCRPFVDGG (SEQ ID NO: 251);GCNIWLNGGDCRAWVDTG (SEQ ID NO: 252); GCNIWIVGGDCRPFINDG (SEQ ID NO:253); GCNIWLNGGDCRPVVFGG (SEQ ID NO: 254); GCNIWLSGGDCRMFMNEG (SEQ IDNO: 255); GCNIWVNGGDCRSFVYSG (SEQ ID NO: 256); GCNIWLNGGDCRGWEASG (SEQID NO: 257); GCNIWAHGGDCRGFIEPG (SEQ ID NO: 258); GCNIWLNGGDCRTFVASG(SEQ ID NO: 259); GCNIWAHGGDCRGFIEPG (SEQ ID NO: 260);GFLENCNIWLNGGDCRTG (SEQ ID NO: 261); GIYENCNIWLNGGDCRMG (SEQ ID NO:262); and/or GIPDNCNIWINGGDCRYG (SEQ ID NO: 263).

Suitable masking moieties for use with antibodies that bind aninterleukin 6 target, e.g., interleukin 6 receptor (IL-6R), include, byway of non-limiting example, masking moieties that include a sequencesuch as QGQSGQYGSCSWNYVHIFMDC (SEQ ID NO: 264); QGQSGQGDFDIPFPAHWVPIT(SEQ ID NO: 265); QGQSGQMGVPAGCVWNYAHIFMDC (SEQ ID NO: 266);YRSCNWNYVSIFLDC (SEQ ID NO: 267); PGAFDIPFPAHWVPNT (SEQ ID NO: 268);ESSCVWNYVHIYMDC (SEQ ID NO: 269); YPGCKWNYDRIFLDC (SEQ ID NO: 270);YRTCSWNYVGIFLDC (SEQ ID NO: 271); YGSCSWNYVHIFMDC (SEQ ID NO: 272);YGSCSWNYVHIFLDC (SEQ ID NO: 273); YGSCNWNYVHIFLDC (SEQ ID NO: 274);YTSCNWNYVHIFMDC (SEQ ID NO: 275); YPGCKWNYDRIFLDC (SEQ ID NO: 276);WRSCNWNYAHIFLDC (SEQ ID NO: 277); WSNCHWNYVHIFLDC (SEQ ID NO: 278);DRSCTWNYVRISYDC (SEQ ID NO: 279); SGSCKWDYVHIFLDC (SEQ ID NO: 280);SRSCIWNYAHIHLDC (SEQ ID NO: 281); SMSCYWQYERIFLDC (SEQ ID NO: 282);YRSCNWNYVSIFLDC (SEQ ID NO: 283); YGSCSWNYVHIFMDC (SEQ ID NO: 284);SGSCKWDYVHIFLDC (SEQ ID NO: 285); YKSCHWDYVHIFLDC (SEQ ID NO: 286);YGSCTWNYVHIFMEC (SEQ ID NO: 287); FSSCNWNYVHIFLDC (SEQ ID NO: 288);WRSCNWNYAHIFLDC (SEQ ID NO: 289); YGSCQWNYVHIFLDC (SEQ ID NO: 290);YRSCNWNYVHIFLDC (SEQ ID NO: 291); NMSCHWDYVHIFLDC (SEQ ID NO: 292);FGPCTWNYARISWDC (SEQ ID NO: 293); XXsCXWXYvhIfXdC (SEQ ID NO: 294);MGVPAGCVWNYAHIFMDC (SEQ ID NO: 295); RDTGGQCRWDYVHIFMDC (SEQ ID NO:296); AGVPAGCTWNYVHIFMEC (SEQ ID NO: 297); VGVPNGCVWNYAHIFMEC (SEQ IDNO: 298); DGGPAGCSWNYVHIFMEC (SEQ ID NO: 299); AVGPAGCWWNYVHIFMEC (SEQID NO: 300); CTWNYVHIFMDCGEGEGP (SEQ ID NO: 301); GGVPEGCTWNYAHIFMEC(SEQ ID NO: 302); AEVPAGCWWNYVHIFMEC (SEQ ID NO: 303);AGVPAGCTWNYVHIFMEC (SEQ ID NO: 304); SGASGGCKWNYVHIFMDC (SEQ ID NO:305); MGVPAGCVWNYAHIFMDC (SEQ ID NO: 306); TPGCRWNYVHIFMECEAL (SEQ IDNO: 307); VGVPNGCVWNYAHIFMEC (SEQ ID NO: 308); PGAFDIPFPAHWVPNT (SEQ IDNO: 309); RGACDIPFPAHWIPNT (SEQ ID NO: 310); QGDFDIPFPAHWVPIT (SEQ IDNO: 311); XGafDIPFPAHWvPnT (SEQ ID NO: 312); RGDGNDSDIPFPAHWVPRT (SEQ IDNO: 313); SGVGRDRDIPFPAHWVPRT (SEQ ID NO: 314); WAGGNDCDIPFPAHWIPNT (SEQID NO: 315); WGDGMDVDIPFPAHWVPVT (SEQ ID NO: 316); AGSGNDSDIPFPAHWVPRT(SEQ ID NO: 317); ESRSGYADIPFPAHWVPRT (SEQ ID NO: 318); and/orRECGRCGDIPFPAHWVPRT (SEQ ID NO: 319).

In some embodiments, the cleavable moiety (CM) of the activatableantibody and/or the multispecific activatable antibody includes an aminoacid sequence that can serve as a substrate for a protease, usually anextracellular protease. The CM may be selected based on a protease thatis produced by a tumor that is in proximity to cells that express thetarget and/or produced by a tumor that is co-localized in tissue withthe desired target of at least one AB of the multispecific activatableantibody. A variety of different conditions are known in which a targetof interest is produced by a tumor that is in proximity to cells thatexpress the target and/or produced by a tumor that is co-localized witha protease, where the substrate of the protease is known in the art. Inthe example of cancer, the target tissue can be a cancerous tissue,particularly cancerous tissue of a solid tumor. There are reports in theliterature of increased levels of proteases having known substrates in anumber of cancers, e.g., solid tumors. See, e.g., La Rocca et al, (2004)British J. of Cancer 90(7): 1414-1421. Non-liming examples of diseaseinclude: all types of cancers (breast, lung, colorectal, prostate,melanomas, head and neck, pancreatic, etc.), rheumatoid arthritis,Crohn's disuse, SLE, cardiovascular damage, ischemia, etc. For example,indications would include leukemias, including T-cell acutelymphoblastic leukemia (T-ALL), lymphoblastic diseases includingmultiple myeloma, and solid tumors, including lung, colorectal,prostate, pancreatic and breast, including triple negative breastcancer. For example, indications include bone disease or metastasis incancer, regardless of primary tumor origin; breast cancer, including byway of non-limiting example, ER/PR+ breast cancer, Her2+ breast cancer,triple-negative breast cancer; colorectal cancer; endometrial cancer;gastric cancer; glioblastoma; head and neck cancer, such as esophagealcancer; lung cancer, such as by way of non-limiting example, non-smallcell lung cancer; multiple myeloma ovarian cancer; pancreatic cancer;prostate cancer; sarcoma, such as osteosarcoma; renal cancer, such as byway of nonlimiting example, renal cell carcinoma; and/or skin cancer,such as by way of nonlimiting example, squamous cell cancer, basal cellcarcinoma, or melanoma. In some embodiments, the cancer is a squamouscell cancer. In some embodiments, the cancer is a skin squamous cellcarcinoma. In some embodiments, the cancer is an esophageal squamouscell carcinoma. In some embodiments, the cancer is a head and necksquamous cell carcinoma. In some embodiments, the cancer is a lungsquamous cell carcinoma.

The CM is specifically cleaved by an enzyme at a rate of about0.001-1500×10⁴ M⁻¹S⁻¹ or at least 0.001, 0.005, 0.01, 0.05, 0.1, 0.5, 1,2.5, 5, 7.5, 10, 15, 20, 25, 50, 75, 100, 125, 150, 200, 250, 500, 750,1000, 1250, or 1500×10⁴ M⁻¹S⁻¹.

For specific cleavage by an enzyme, contact between the enzyme and CM ismade. When the activatable antibody comprising an AB coupled to a MM anda CM is in the presence of target and sufficient enzyme activity, the CMcan be cleaved. When the multispecific activatable antibody comprisingat least a first AB coupled to a MM and a CM is in the presence oftarget and sufficient enzyme activity, the CM can be cleaved. Sufficientenzyme activity can refer to the ability of the enzyme to make contactwith the CM and effect cleavage. It can readily be envisioned that anenzyme may be in the vicinity of the CM but unable to cleave because ofother cellular factors or protein modification of the enzyme.

Exemplary substrates include but are not limited to substrates cleavableby one or more of the following enzymes or proteases in Table 3:

TABLE 3 Exemplary Proteases and/or Enzymes ADAMS, ADAMTS, e.g. ADAM8ADAM9 ADAM10 ADAM12 ADAM15 ADAM17/TACE ADAMDEC1 ADAMTS1 ADAMTS4 ADAMTS5Aspartate proteases, e.g., BACE Renin Aspartic cathepsins, e.g.,Cathepsin D Cathepsin E Caspases, e.g., Caspase 1 Caspase 2 Caspase 3Caspase 4 Caspase 5 Caspase 6 Caspase 7 Caspase 8 Caspase 9 Caspase 10Caspase 14 Cysteine cathepsins, e.g., Cathepsin B Cathepsin C CathepsinK Cathepsin L Cathepsin S Cathepsin V/L2 Cathepsin X/Z/P Cysteineproteinases, e.g., Cruzipain Legumain Otubain-2 KLKs, e.g., KLK4 KLK5KLK6 KLK7 KLK8 KLK10 KLK11 KLK13 KLK14 Metallo proteinases, e.g., MeprinNeprilysin PSMA BMP-1 MMPs, e.g., MMP1 MMP2 MMP3 MMP7 MMP8 MMP9 MMP10MMP11 MMP12 MMP13 MMP14 MMP15 MMP16 MMP17 MMP19 MMP20 MMP23 MMP24 MMP26MMP27 Serine proteases, e.g., activated protein C Cathepsin A CathepsinG Chymase coagulation factor proteases (e.g., FVIIa, FIXa, FXa, FXIa,FXIIa) Elastase Granzyme B Guanidinobenzoatase HtrA1 Human NeutrophilElastase Lactoferrin Marapsin NS3/4A PACE4 Plasmin PSA tPA ThrombinTryptase uPA Type II Transmembrane Serine Proteases (TTSPs), e.g., DESC1DPP-4 FAP Hepsin Matriptase-2 Matriptase TMPRSS2 TMRSS3 TMPRSS4

For example, in some embodiments, the substrate is cleavable by one ormore of the following enzymes or proteases: uPA, legumain, matriptase,ADAM17, BMP-1, TMPRSS3, TMPRSS4, MMP-9, MMP-12, MMP-13, and/or MMP-14.In some embodiments, the protease is selected from the group of uPA,legumain, and matriptase. In some embodiments, the protease is a matrixmetalloproteinase. In some embodiments, the protease comprises uPA. Insome embodiments, the protease comprises legumain. In some embodiments,the protease comprises matriptase.

In some embodiments, the CM is selected for use with a specificprotease. In some embodiments, the CM is a substrate for at least oneprotease selected from the group consisting of an ADAM 17, a BMP-1, acysteine protease such as a cathepsin, a HtrA1, a legumain, a matriptase(MT-SP1), a matrix metalloprotease (MMP), a neutrophil elastase, aTMPRSS, such as TMPRSS3 or TMPRSS4, a thrombin, and a u-type plasminogenactivator (uPA, also referred to as urokinase).

In some embodiments, the CM is a substrate for an ADAM17. In someembodiments, the CM is a substrate for a BMP-1. In some embodiments, theCM is a substrate for a cathepsin. In some embodiments, the CM is asubstrate for a cysteine protease. In some embodiments, the CM is asubstrate for a HtrA1. In some embodiments, the CM is a substrate for alegumain. In some embodiments, the CM is a substrate for matriptase. Insome embodiments, the CM is a substrate for a MMP. In some embodiments,the CM is a substrate for a neutrophil elastase. In some embodiments,the CM is a substrate for a thrombin. In some embodiments, the CM is asubstrate for a TMPRSS. In some embodiments, the CM is a substrate forTMPRSS3. In some embodiments, the CM is a substrate for TMPRSS4. In someembodiments, the CM is a substrate for uPA.

In some embodiments, the cleavable moiety is selected for use with aspecific protease, for example a protease that is known to be producedby a tumor that is in proximity to cells that express the target and/orproduced by a tumor that is co-localized with the target of theactivatable antibody. For example, suitable cleavable moieties for usein the activatable antibodies of the disclosure include the sequenceTGRGPSWV (SEQ ID NO: 68); SARGPSRW (SEQ ID NO: 69); TARGPSFK (SEQ ID NO:70); LSGRSDNH (SEQ ID NO: 67); GGWHTGRN (SEQ ID NO: 71); HTGRSGAL (SEQID NO: 72); PLTGRSGG (SEQ ID NO: 73); AARGPAIH (SEQ ID NO: 74); RGPAFNPM(SEQ ID NO: 75); SSRGPAYL (SEQ ID NO: 76); RGPATPIM (SEQ ID NO: 77);RGPA (SEQ ID NO: 78); GGQPSGMWGW (SEQ ID NO: 79); FPRPLGITGL (SEQ ID NO:80); VHMPLGFLGP (SEQ ID NO: 81); SPLTGRSG (SEQ ID NO: 82); SAGFSLPA (SEQID NO: 83); LAPLGLQRR (SEQ ID NO: 84); SGGPLGVR (SEQ ID NO: 85); and/orPLGL (SEQ ID NO: 86).

In some embodiments, the CM is a substrate for at least one matrixmetalloprotease (MMP). Examples of MMPs include MMP1; MMP2; MMP3; MMP7;MMP8; MMP9; MMP10; MMP11; MMP12; MMP13; MMP14; MMP15; MMP16; MMP17;MMP19; MMP20; MMP23; MMP24; MMP26; and MMP27. In some embodiments, theCM is a substrate for MMP9, MMP14, MMP1, MMP3, MMP13, MMP17, MMP11, andMMP19. In some embodiments, the CM is a substrate for MMP7. In someembodiments, the CM is a substrate for MMP9. In some embodiments, the CMis a substrate for MMP14. In some embodiments, the CM is a substrate fortwo or more MMPs. In some embodiments, the CM is a substrate for atleast MMP9 and MMP14. In some embodiments, the CM comprises two or moresubstrates for the same MMP. In some embodiments, the CM comprises atleast two or more MMP9 substrates. In some embodiments, the CM comprisesat least two or more MMP14 substrates.

In some embodiments, the CM is a substrate for an MMP and includes thesequence ISSGLLSS (SEQ ID NO: 321); QNQALRMA (SEQ ID NO: 322); AQNLLGMV(SEQ ID NO: 323); STFPFGMF (SEQ ID NO: 324); PVGYTSSL (SEQ ID NO: 325);DWLYWPGI (SEQ ID NO: 326); MIAPVAYR (SEQ ID NO: 327); RPSPMWAY (SEQ IDNO: 328); WATPRPMR (SEQ ID NO: 329); FRLLDWQW (SEQ ID NO: 330); LKAAPRWA(SEQ ID NO: 331); GPSHLVLT (SEQ ID NO: 332); LPGGLSPW (SEQ ID NO: 333);MGLFSEAG (SEQ ID NO: 334); SPLPLRVP (SEQ ID NO: 335); RMHLRSLG (SEQ IDNO: 336); LAAPLGLL (SEQ ID NO: 337); AVGLLAPP (SEQ ID NO: 338); LLAPSHRA(SEQ ID NO: 339); PAGLWLDP (SEQ ID NO: 340); and/or ISSGLSS (SEQ ID NO:341).

In some embodiments, the CM is a substrate for thrombin. In someembodiments, the CM is a substrate for thrombin and includes thesequence GPRSFGL (SEQ ID NO: 896) or GPRSFG (SEQ ID NO: 897).

In some embodiments, the CM comprises an amino acid sequence selectedfrom the group consisting of NTLSGRSENHSG (SEQ ID NO: 898); NTLSGRSGNHGS(SEQ ID NO: 899); TSTSGRSANPRG (SEQ ID NO: 900); TSGRSANP (SEQ ID NO:901); VAGRSMRP (SEQ ID NO: 902); VVPEGRRS (SEQ ID NO: 903); ILPRSPAF(SEQ ID NO: 904); MVLGRSLL (SEQ ID NO: 905); QGRAITFI (SEQ ID NO: 906);SPRSIMLA (SEQ ID NO: 907); and SMLRSMPL (SEQ ID NO: 908).

In some embodiments, the CM is a substrate for a neutrophil elastase. Insome embodiments, the CM is a substrate for a serine protease. In someembodiments, the CM is a substrate for uPA. In some embodiments, the CMis a substrate for legumain. In some embodiments, the CM is a substratefor matriptase. In some embodiments, the CM is a substrate for acysteine protease. In some embodiments, the CM is a substrate for acysteine protease, such as a cathepsin.

In some embodiments, the CM is a CM1-CM2 substrate and includes thesequence ISSGLLSGRSDNH (SEQ ID NO: 909); ISSGLLSSGGSGGSLSGRSDNH (SEQ IDNO: 910); AVGLLAPPGGTSTSGRSANPRG (SEQ ID NO: 911);TSTSGRSANPRGGGAVGLLAPP (SEQ ID NO: 912); VHMPLGFLGPGGTSTSGRSANPRG (SEQID NO: 913); TSTSGRSANPRGGGVHMPLGFLGP (SEQ ID NO: 914);AVGLLAPPGGLSGRSDNH (SEQ ID NO: 915); LSGRSDNHGGAVGLLAPP (SEQ ID NO:916); VHMPLGFLGPGGLSGRSDNH (SEQ ID NO: 917); LSGRSDNHGGVHMPLGFLGP (SEQID NO: 918); LSGRSDNHGGSGGSISSGLLSS (SEQ ID NO: 919);LSGRSGNHGGSGGSISSGLLSS (SEQ ID NO: 920); ISSGLLSSGGSGGSLSGRSGNH (SEQ IDNO: 921); LSGRSDNHGGSGGSQNQALRMA (SEQ ID NO: 922);QNQALRMAGGSGGSLSGRSDNH (SEQ ID NO: 923); LSGRSGNHGGSGGSQNQALRMA (SEQ IDNO: 924); QNQALRMAGGSGGSLSGRSGNH (SEQ ID NO: 925) and/or ISSGLLSGRSGNH(SEQ ID NO: 926).

In some embodiments, anti-CD3ε antibodies, activatable antibodies and/ormultispecific antibodies and/or multispecific activatable antibodies ofthe disclosure may be made biosynthetically using recombinant DNAtechnology and expression in eukaryotic or prokaryotic species. For theactivatable antibodies and/or multispecific activatable antibodies, thecDNAs encoding the masking moiety, linker sequence (that may include acleavable moiety (CM), and antibody chain (heavy or light)) can belinked in an 5′ to 3′ (N- to C-terminal in the translated product)sequence to create the nucleic acid construct, which is expressed as theactivatable antibody protein and/or multispecific activatable antibodyprotein following a conventional antibody expression process. In someembodiments, the activatable antibody and/or multispecific activatableantibody could be semi-synthetically produced by expressing aCM-antibody and then coupling the mask chemically at or near theN-terminus of the protein. In some embodiments, the activatable antibodyand/or the multispecific activatable antibody could be produced byexpressing an antibody and then coupling the mask and the CM chemicallyat or near the N-terminus of the protein such that the activatableantibody and/or at least a portion of the multispecific activatableantibody in the uncleaved state has the structural arrangement fromN-terminus to C-terminus as follows: MM-CM-AB or AB-CM-MM.

Linkers suitable for use in compositions described herein are generallyones that provide flexibility of the modified AB or the multispecificactivatable antibodies to facilitate the inhibition of the binding of atleast the first AB to the target. Such linkers are generally referred toas flexible linkers. Suitable linkers can be readily selected and can beof any of a suitable of different lengths, such as from 1 amino acid(e.g., Gly) to 20 amino acids, from 2 amino acids to 15 amino acids,from 3 amino acids to 12 amino acids, including 4 amino acids to 10amino acids, 5 amino acids to 9 amino acids, 6 amino acids to 8 aminoacids, or 7 amino acids to 8 amino acids, and may be 1, 2, 3, 4, 5, 6,7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 amino acids inlength.

Exemplary flexible linkers include glycine polymers (G)n, glycine-serinepolymers (including, for example, (GS)n, (GSGGS)n (SEQ ID NO: 59) and(GGGS)n (SEQ ID NO: 60), where n is an integer of at least one),glycine-alanine polymers, alanine-serine polymers, and other flexiblelinkers known in the art. Glycine and glycine-serine polymers arerelatively unstructured, and therefore may be able to serve as a neutraltether between components. Glycine accesses significantly more phi-psispace than even alanine, and is much less restricted than residues withlonger side chains (see Scheraga, Rev. Computational Chem. 11173-142(1992)). Exemplary flexible linkers include, but are not limited to GGSG(SEQ ID NO: 61), GGSGG(SEQ ID NO: 62), GSGSG(SEQ ID NO: 63), GSGGG(SEQID NO: 64), GGGSG(SEQ ID NO: 65), GSSSG (SEQ ID NO: 66), and the like.The ordinarily skilled artisan will recognize that design of anactivatable antibodies can include linkers that are all or partiallyflexible, such that the linker can include a flexible linker as well asone or more portions that confer less flexible structure to provide fora desired activatable antibodies and/or multispecific activatableantibodies structure.

In addition to the elements described above, the activatable antibodiesand/or multispecific activatable antibodies can contain additionalelements such as, for example, amino acid sequence N- or C-terminal ofthe activatable antibodies and/or multispecific activatable antibodies.For example, activatable antibodies and/or multispecific activatableantibodies can include a targeting moiety to facilitate delivery to acell or tissue of interest. Activatable antibodies and/or multispecificactivatable antibodies can be conjugated to an agent, such as atherapeutic agent, a detectable moiety or a diagnostic agent. Examplesof agents are disclosed herein.

The activatable antibodies and/or multispecific activatable antibodiescan also include any of the conjugated agents, linkers and othercomponents described herein in conjunction with an activatable antibodyand/or a multispecific antibody of the disclosure, including by way ofnon-limiting example, any of the agents listed in Table 4 and/or any ofthe linkers listed in Table 5 and/or Table 6.

Collimated Antibodies, Collimated Activatable Antibodies, CollimatedMultispecific Antibodies and Collimated Multispecific ActivatableAntibodies

In some embodiments, the antibodies and/or activatable antibodies of thedisclosure can be conjugated to an agent, such as for example, atherapeutic agent, a detectable moiety or a diagnostic agent. Suitabletherapeutic agents include, by way of non-limiting example, atherapeutic agent for use in targeting a T cell-derived lymphoma.Suitable detectable agents include, by way of non-limiting example, afluorescent dye, (e.g. a fluorophore, Fluorescein Isothiocyanate (FITC),Rhodamine Isothiocyanate (TRITC), an Alexa Fluor® label), a nearinfrared (NIR) dye (e.g., Qdot® nanocrystals), a colloidal metal, ahapten, a radioactive marker, biotin and an amplification reagent suchas streptavidin, or an enzyme (e.g. horseradish peroxidase or alkalinephosphatase).

The activatable anti-CD3ε antibodies have at least one point ofconjugation for an agent, but in the methods and compositions providedherein less than all possible points of conjugation are available forconjugation to an agent. In some embodiments, the one or more points ofconjugation are sulfur atoms involved in disulfide bonds. In someembodiments, the one or more points of conjugation are sulfur atomsinvolved in interchain disulfide bonds. In some embodiments, the one ormore points of conjugation are sulfur atoms involved in interchainsulfide bonds, but not sulfur atoms involved in intrachain disulfidebonds. In some embodiments, the one or more points of conjugation aresulfur atoms of cysteine or other amino acid residues containing asulfur atom. Such residues may occur naturally in the antibody structureor may be incorporated into the antibody by site-directed mutagenesis,chemical conversion, or mis-incorporation of non-natural amino acids.

Also provided are methods of preparing a conjugate of an activatableanti-CD3ε antibody having one or more interchain disulfide bonds in theAB and one or more intrachain disulfide bonds in the MM, and a drugreactive with free thiols is provided. The method generally includespartially reducing interchain disulfide bonds in the activatableantibody with a reducing agent, such as, for example, TCEP; andconjugating the drug reactive with free thiols to the partially reducedactivatable antibody. As used herein, the term partial reduction refersto situations where an activatable anti-CD3ε antibody is contacted witha reducing agent and less than all disulfide bonds, e.g., less than allpossible sites of conjugation are reduced. In some embodiments, lessthan 99%, 98%, 97%, 96%, 95%, 90%, 85%, 80%, 75%, 70%, 65%, 60%, 55%,50%, 45%, 40%, 35%, 30%, 25%, 20%, 15%, 10% or less than 5% of allpossible sites of conjugation are reduced.

In yet other embodiments, a method of reducing and conjugating an agent,e.g., a drug, to an activatable anti-CD3ε antibody resulting inselectivity in the placement of the agent is provided. The methodgenerally includes partially reducing the activatable anti-CD3ε antibodywith a reducing agent such that any conjugation sites in the maskingmoiety or other non-AB portion of the activatable antibody are notreduced, and conjugating the agent to interchain thiols in the AB. Theconjugation site(s) are selected so as to allow desired placement of anagent to allow conjugation to occur at a desired site. The reducingagent is, for example, TCEP. The reduction reaction conditions such as,for example, the ratio of reducing agent to activatable antibody, thelength of incubation, the temperature during the incubation, the pH ofthe reducing reaction solution, etc., are determined by identifying theconditions that produce a conjugated activatable antibody in which theMM retains the ability to effectively and efficiently mask the AB of theactivatable antibody in an uncleaved state. The ratio of reduction agentto activatable anti-CD3ε antibody will vary depending on the activatableantibody. In some embodiments, the ratio of reducing agent toactivatable anti-CD3ε antibody will be in a range from about 20:1 to1:1, from about 10:1 to 1:1, from about 9:1 to 1:1, from about 8:1 to1:1, from about 7:1 to 1:1, from about 6:1 to 1:1, from about 5:1 to1:1, from about 4:1 to 1:1, from about 3:1 to 1:1, from about 2:1 to1:1, from about 20:1 to 1:1.5, from about 10:1 to 1:1.5, from about 9:1to 1:1.5, from about 8:1 to 1:1.5, from about 7:1 to 1:1.5, from about6:1 to 1:1.5, from about 5:1 to 1:1.5, from about 4:1 to 1:1.5, fromabout 3:1 to 1:1.5, from about 2:1 to 1:1.5, from about 1.5:1 to 1:1.5,or from about 1:1 to 1:1.5. In some embodiments, the ratio is in a rangeof from about 5:1 to 1:1. In some embodiments, the ratio is in a rangeof from about 5:1 to 1.5:1. In some embodiments, the ratio is in a rangeof from about 4:1 to 1:1. In some embodiments, the ratio is in a rangefrom about 4:1 to 1.5:1. In some embodiments, the ratio is in a rangefrom about 8:1 to about 1:1. In some embodiments, the ratio is in arange of from about 2.5:1 to 1:1.

In some embodiments, a method of reducing interchain disulfide bonds inthe AB of an activatable anti-CD3ε antibody and conjugating an agent,e.g., a thiol-containing agent such as a drug, to the resultinginterchain thiols to selectively locate agent(s) on the AB is provided.The method generally includes partially reducing the AB with a reducingagent to form at least two interchain thiols without forming allpossible interchain thiols in the activatable antibody; and conjugatingthe agent to the interchain thiols of the partially reduced AB. Forexample, the AB of the activatable antibody is partially reduced forabout 1 hour at about 37° C. at a desired ratio of reducingagent:activatable antibody. In some embodiments, the ratio of reducingagent to activatable antibody will be in a range from about 20:1 to 1:1,from about 10:1 to 1:1, from about 9:1 to 1:1, from about 8:1 to 1:1,from about 7:1 to 1:1, from about 6:1 to 1:1, from about 5:1 to 1:1,from about 4:1 to 1:1, from about 3:1 to 1:1, from about 2:1 to 1:1,from about 20:1 to 1:1.5, from about 10:1 to 1:1.5, from about 9:1 to1:1.5, from about 8:1 to 1:1.5, from about 7:1 to 1:1.5, from about 6:1to 1:1.5, from about 5:1 to 1:1.5, from about 4:1 to 1:1.5, from about3:1 to 1:1.5, from about 2:1 to 1:1.5, from about 1.5:1 to 1:1.5, orfrom about 1:1 to 1:1.5. In some embodiments, the ratio is in a range offrom about 5:1 to 1:1. In some embodiments, the ratio is in a range offrom about 5:1 to 1.5:1. In some embodiments, the ratio is in a range offrom about 4:1 to 1:1. In some embodiments, the ratio is in a range fromabout 4:1 to 1.5:1. In some embodiments, the ratio is in a range fromabout 8:1 to about 1:1. In some embodiments, the ratio is in a range offrom about 2.5:1 to 1:1.

The thiol-containing reagent can be, for example, cysteine or N-acetylcysteine. The reducing agent can be, for example, TCEP. In someembodiments, the reduced activatable antibody can be purified prior toconjugation, using for example, column chromatography, dialysis, ordiafiltration. In some embodiments, the reduced antibody is not purifiedafter partial reduction and prior to conjugation.

The disclosure also provides partially reduced activatable anti-CD3εantibodies in which at least one interchain disulfide bond in theactivatable antibody has been reduced with a reducing agent withoutdisturbing any intrachain disulfide bonds in the activatable antibody,wherein the activatable antibody includes an antibody or an antigenbinding fragment thereof (AB) that specifically binds to a CD3ε target,a masking moiety (MM) that inhibits the binding of the AB of theactivatable antibody in an uncleaved state to the CD3ε target, and acleavable moiety (CM) coupled to the AB, wherein the CM is a polypeptidethat functions as a substrate for a protease. In some embodiments the MMis coupled to the AB via the CM. In some embodiments, one or moreintrachain disulfide bond(s) of the activatable antibody is notdisturbed by the reducing agent. In some embodiments, one or moreintrachain disulfide bond(s) of the MM within the activatable antibodyis not disturbed by the reducing agent. In some embodiments, theactivatable antibody in the uncleaved state has the structuralarrangement from N-terminus to C-terminus as follows: MM-CM-AB orAB-CM-MM. In some embodiments, reducing agent is TCEP.

The compositions and methods provided herein enable the attachment ofone or more agents to one or more cysteine residues in any of the ABregions without compromising the activity (e.g., the masking, activatingor binding activity) of the multispecific activatable antibody. In someembodiments, the compositions and methods provided herein enable theattachment of one or more agents to one or more cysteine residues in anyof the AB regions without reducing or otherwise disturbing one or moredisulfide bonds within any of the MM. The compositions and methodsprovided herein produce a multispecific activatable antibody that isconjugated to one or more agents, e.g., any of a variety of therapeutic,diagnostic and/or prophylactic agents, preferably without any of theagent(s) being conjugated to any of the MM of the multispecificactivatable antibody. The compositions and methods provided hereinproduce conjugated multispecific activatable antibodies in which each ofthe MM retains the ability to effectively and efficiently mask itscorresponding AB of the multispecific activatable antibody in anuncleaved state. The compositions and methods provided herein produceconjugated multispecific activatable antibodies in which the activatableantibody is still activated, i.e., cleaved, in the presence of aprotease that can cleave the CM.

The multispecific activatable antibodies have at least one point ofconjugation for an agent, but in the methods and compositions providedherein less than all possible points of conjugation are available forconjugation to an agent. In some embodiments, the one or more points ofconjugation are sulfur atoms involved in disulfide bonds. In someembodiments, the one or more points of conjugation are sulfur atomsinvolved in interchain disulfide bonds. In some embodiments, the one ormore points of conjugation are sulfur atoms involved in interchainsulfide bonds, but not sulfur atoms involved in intrachain disulfidebonds. In some embodiments, the one or more points of conjugation aresulfur atoms of cysteine or other amino acid residues containing asulfur atom. Such residues may occur naturally in the antibody structureor may be incorporated into the antibody by site-directed mutagenesis,chemical conversion, or mis-incorporation of non-natural amino acids.

Also provided are methods of preparing a conjugate of a multispecificactivatable antibody having one or more interchain disulfide bonds inone or more of the AB and one or more intrachain disulfide bonds in thecorresponding MM, and a drug reactive with free thiols is provided. Themethod generally includes partially reducing interchain disulfide bondsin the activatable antibody with a reducing agent, such as, for example,TCEP; and conjugating the drug reactive with free thiols to thepartially reduced activatable antibody. As used herein, the term partialreduction refers to situations where a multispecific activatableantibody is contacted with a reducing agent and less than all disulfidebonds, e.g., less than all possible sites of conjugation are reduced. Insome embodiments, less than 99%, 98%, 97%, 96%, 95%, 90%, 85%, 80%, 75%,70%, 65%, 60%, 55%, 50%, 45%, 40%, 35%, 30%, 25%, 20%, 15%, 10% or lessthan 5% of all possible sites of conjugation are reduced.

In yet other embodiments, a method of reducing and conjugating an agent,e.g., a drug, to a multispecific activatable antibody resulting inselectivity in the placement of the agent is provided. The methodgenerally includes partially reducing the multispecific activatableantibody with a reducing agent such that any conjugation sites in any ofthe masking moieties or other non-AB portion of the activatable antibodyare not reduced, and conjugating the agent to interchain thiols in oneor more of the AB regions of the multispecific activatable antibody. Theconjugation site(s) are selected so as to allow desired placement of anagent to allow conjugation to occur at a desired site. The reducingagent is, for example, TCEP. The reduction reaction conditions such as,for example, the ratio of reducing agent to activatable antibody, thelength of incubation, the temperature during the incubation, the pH ofthe reducing reaction solution, etc., are determined by identifying theconditions that produce a conjugated activatable antibody in which theMM retains the ability to effectively and efficiently mask the AB of theactivatable antibody in an uncleaved state. The ratio of reduction agentto multispecific activatable antibody will vary depending on theactivatable antibody. In some embodiments, the ratio of reducing agentto multispecific activatable antibody will be in a range from about 20:1to 1:1, from about 10:1 to 1:1, from about 9:1 to 1:1, from about 8:1 to1:1, from about 7:1 to 1:1, from about 6:1 to 1:1, from about 5:1 to1:1, from about 4:1 to 1:1, from about 3:1 to 1:1, from about 2:1 to1:1, from about 20:1 to 1:1.5, from about 10:1 to 1:1.5, from about 9:1to 1:1.5, from about 8:1 to 1:1.5, from about 7:1 to 1:1.5, from about6:1 to 1:1.5, from about 5:1 to 1:1.5, from about 4:1 to 1:1.5, fromabout 3:1 to 1:1.5, from about 2:1 to 1:1.5, from about 1.5:1 to 1:1.5,or from about 1:1 to 1:1.5. In some embodiments, the ratio is in a rangeof from about 5:1 to 1:1. In some embodiments, the ratio is in a rangeof from about 5:1 to 1.5:1. In some embodiments, the ratio is in a rangeof from about 4:1 to 1:1. In some embodiments, the ratio is in a rangefrom about 4:1 to 1.5:1. In some embodiments, the ratio is in a rangefrom about 8:1 to about 1:1. In some embodiments, the ratio is in arange of from about 2.5:1 to 1:1.

In some embodiments, a method of reducing interchain disulfide bonds inone or more of the AB regions of a multispecific activatable antibodyand conjugating an agent, e.g., a thiol-containing agent such as a drug,to the resulting interchain thiols to selectively locate agent(s) on theAB is provided. The method generally includes partially reducing one ormore of the AB regions with a reducing agent to form at least twointerchain thiols without forming all possible interchain thiols in theactivatable antibody; and conjugating the agent to the interchain thiolsof the partially reduced AB. For example, one or more of the AB regionsof the multispecific activatable antibody is partially reduced for about1 hour at about 37° C. at a desired ratio of reducing agent:activatableantibody. In some embodiments, the ratio of reducing agent toactivatable antibody will be in a range from about 20:1 to 1:1, fromabout 10:1 to 1:1, from about 9:1 to 1:1, from about 8:1 to 1:1, fromabout 7:1 to 1:1, from about 6:1 to 1:1, from about 5:1 to 1:1, fromabout 4:1 to 1:1, from about 3:1 to 1:1, from about 2:1 to 1:1, fromabout 20:1 to 1:1.5, from about 10:1 to 1:1.5, from about 9:1 to 1:1.5,from about 8:1 to 1:1.5, from about 7:1 to 1:1.5, from about 6:1 to1:1.5, from about 5:1 to 1:1.5, from about 4:1 to 1:1.5, from about 3:1to 1:1.5, from about 2:1 to 1:1.5, from about 1.5:1 to 1:1.5, or fromabout 1:1 to 1:1.5. In some embodiments, the ratio is in a range of fromabout 5:1 to 1:1. In some embodiments, the ratio is in a range of fromabout 5:1 to 1.5:1. In some embodiments, the ratio is in a range of fromabout 4:1 to 1:1. In some embodiments, the ratio is in a range fromabout 4:1 to 1.5:1. In some embodiments, the ratio is in a range fromabout 8:1 to about 1:1. In some embodiments, the ratio is in a range offrom about 2.5:1 to 1:1.

The thiol-containing reagent can be, for example, cysteine or N-acetylcysteine. The reducing agent can be, for example, TCEP. In someembodiments, the reduced activatable antibody can be purified prior toconjugation, using for example, column chromatography, dialysis, ordiafiltration. In some embodiments, the reduced antibody is not purifiedafter partial reduction and prior to conjugation.

The disclosure also provides partially reduced multispecific activatableantibodies in which at least one interchain disulfide bond in themultispecific activatable antibody has been reduced with a reducingagent without disturbing any intrachain disulfide bonds in themultispecific activatable antibody, wherein the multispecificactivatable antibody includes at least a first antibody or an antigenbinding fragment thereof (AB1) that specifically binds to a target, afirst masking moiety (MM1) that inhibits the binding of the AB1 of themultispecific activatable antibody in an uncleaved state to the target,a first cleavable moiety (CM1) coupled to the AB1, wherein the CM1 is apolypeptide that functions as a substrate for a protease, and a secondantibody or an antigen binding fragment thereof (AB2) that specificallybinds to a second target. In some embodiments, the MM1 is coupled to theAB1 via the CM1. In some embodiments, one or more intrachain disulfidebond(s) of the multispecific activatable antibody is not disturbed bythe reducing agent. In some embodiments, one or more intrachaindisulfide bond(s) of the MM1 within the multispecific activatableantibody is not disturbed by the reducing agent. In some embodiments,reducing agent is TCEP.

The disclosure also pertains to immunoconjugates comprising an anti-CD3εantibody, an activatable antibody, a multispecific antibody and/or amultispecific activatable antibody conjugated to a cytotoxic agent suchas a toxin (e.g., an enzymatically active toxin of bacterial, fungal,plant, or animal origin, or fragments thereof), or a radioactive isotope(i.e., a radioconjugate). Suitable cytotoxic agents for use in targetingdiseased T cells such as in a T cell-derived lymphoma include, forexample, dolastatins and derivatives thereof (e.g. auristatin E, AFP,MMAD, MMAF, MMAE). For example, the cytotoxic agent is monomethylauristatin E (MMAE). In some embodiments, the agent is monomethylauristatin E (MMAD). In some embodiments, the agent is an agent selectedfrom the group listed in Table 4. In some embodiments, the agent is adolastatin. In some embodiments, the agent is an auristatin orderivative thereof. In some embodiments, the agent is auristatin E or aderivative thereof. In some embodiments, the agent is monomethylauristatin E (MMAE). In some embodiments, the agent is monomethylauristatin D (MMAD). In some embodiments, the agent is a maytansinoid ormaytansinoid derivative. In some embodiments, the agent is DM1 or DM4.In some embodiments, the agent is a duocarmycin or derivative thereof.In some embodiments, the agent is a calicheamicin or derivative thereof.In some embodiments, the agent is a pyrrolobenzodiazepine.

Enzymatically active toxins and fragments thereof that can be usedinclude diphtheria A chain, nonbinding active fragments of diphtheriatoxin, exotoxin A chain (from Pseudomonas aeruginosa), ricin A chain,abrin A chain, modeccin A chain, alpha-sarcin, Aleurites fordiiproteins, dianthin proteins, Phytolaca americana proteins (PAPI, PAPII,and PAP-S), momordica charantia inhibitor, curcin, crotin, sapaonariaofficinalis inhibitor, gelonin, mitogellin, restrictocin, phenomycin,enomycin, and the tricothecenes. A variety of radionuclides areavailable for the production of radioconjugated antibodies. Examplesinclude ²¹²Bi, ⁶⁴Cu, ¹²⁵I, ¹³¹I, ¹³¹In, ^(99m)Tc, ⁹⁰Y, ¹⁸⁶Re, and ⁸⁹Zr.

Conjugates of the antibody and cytotoxic agent are made using a varietyof bifunctional protein-coupling agents such asN-succinimidyl-3-(2-pyridyldithiol) propionate (SPDP), iminothiolane(IT), bifunctional derivatives of imidoesters (such as dimethyladipimidate HCL), active esters (such as disuccinimidyl suberate),aldehydes (such as glutaraldehyde), bis-azido compounds (such as bis(p-azidobenzoyl) hexanediamine), bis-diazonium derivatives (such asbis-(p-diazoniumbenzoyl)-ethylenediamine), diisocyanates (such astolyene 2,6-diisocyanate), and bis-active fluorine compounds (such as1,5-difluoro-2,4-dinitrobenzene). For example, a ricin immunotoxin canbe prepared as described in Vitetta et al., Science 238: 1098 (1987).Carbon-14-labeled 1-isothiocyanatobenzyl-3-methyldiethylenetriaminepentaacetic acid (MX-DTPA) is an exemplary chelating agent forconjugation of radionucleotide to the antibody. (See WO94/11026).

Table 4 lists some of the exemplary pharmaceutical agents that may beemployed in the herein described disclosure but in no way is meant to bean exhaustive list.

TABLE 4 Exemplary Pharmaceutical Agents for Conjugation CYTOTOXIC AGENTSAuristatins Auristatin E Monomethyl auristatin D (MMAD) Monomethylauristatin E (MMAE) Desmethyl auristatin E (DMAE) Auristatin FMonomethyl auristatin F (MMAF) Desmethyl auristatin F (DMAF) Auristatinderivatives, e.g., amides thereof Auristatin tyramine Auristatinquinolone Dolastatins Dolastatin derivatives Dolastatin 16 DmJDolastatin 16 Dpv Maytansinoids, e.g. DM-1; DM-4 Maytansinoidderivatives Duocarmycin Duocarmycin derivatives Alpha-amanitinAnthracyclines Doxorubicin Daunorubicin Bryostatins CamptothecinCamptothecin derivatives 7-substituted Camptothecin 10,11-Difluoromethylenedioxycamptothecin Combretastatins DebromoaplysiatoxinKahalalide-F Discodermolide Ecteinascidins ANTIVIRALS Acyclovir Vira ASymmetrel ANTIFUNGALS Nystatin ADDITIONAL ANTI-NEOPLASTICS AdriamycinCerubidine Bleomycin Alkeran Velban Oncovin Fluorouracil MethotrexateThiotepa Bisantrene Novantrone Thioguanine Procarabizine CytarabineANTI-BACTERIALS Aminoglycosides Streptomycin Neomycin Kanamycin AmikacinGentamicin Tobramycin Streptomycin B Spectinomycin AmpicillinSulfanilamide Polymyxin Chloramphenicol Turbostatin PhenstatinsHydroxyphenstatin Spongistatin 5 Spongistatin 7 Halistatin 1 Halistatin2 Halistatin 3 Modified Bryostatins Halocomstatins Pyrrolobenzimidazoles(PBI) Cibrostatin6 Doxaliform Anthracyclins analogues Cemadotin analogue(CemCH2-SH) Pseudomonas toxin A (PE38) variant Pseudomonas toxin A(ZZ-PE38) variant ZJ-101 OSW-1 4-Nitrobenzyloxycarbonyl Derivatives ofO6-Benzylguanine Topoisomerase inhibitors Hemiasterlin CephalotaxineHomoharringtonine Pyrrolobenzodiazepine dimers (PBDs) Functionalizedpyrrolobenzodiazepenes Calicheamicins Podophyllotoxins Taxanes Vincaalkaloids CONJUGATABLE DETECTION REAGENTS Fluorescein and derivativesthereof Fluorescein isothiocyanate (FITC) RADIOPHARMACEUTICALS ¹²⁵I ¹³¹I⁸⁹Zr ¹¹¹In ¹²³I ¹³¹I ⁹⁹mTc ²⁰¹Tl ¹³³Xe ¹¹C ⁶²Cu ¹⁸F ⁶⁸Ga ¹³N ¹⁵O ³⁸K⁸²Rb ⁹⁹mTc (Technetium) HEAVY METALS Barium Gold PlatinumANTI-MYCOPLASMALS Tylosine Spectinomycin

Those of ordinary skill in the art will recognize that a large varietyof possible moieties can be coupled to the resultant anti-CD3εantibodies, activatable antibodies, multispecific antibodies and/ormultispecific activatable antibodies of the disclosure. (See, forexample, “Conjugate Vaccines”, Contributions to Microbiology andImmunology, J. M. Cruse and R. E. Lewis, Jr (eds), Carger Press, NewYork, (1989), the entire contents of which are incorporated herein byreference).

Coupling may be accomplished by any chemical reaction that will bind thetwo molecules so long as the antibody and the other moiety retain theirrespective activities. This linkage can include many chemicalmechanisms, for instance covalent binding, affinity binding,intercalation, coordinate binding and complexation. In some embodiments,the preferred binding is, however, covalent binding. Covalent bindingcan be achieved either by direct condensation of existing side chains orby the incorporation of external bridging molecules. Many bivalent orpolyvalent linking agents are useful in coupling protein molecules, suchas the antibodies of the present disclosure, to other molecules. Forexample, representative coupling agents can include organic compoundssuch as thioesters, carbodiimides, succinimide esters, diisocyanates,glutaraldehyde, diazobenzenes and hexamethylene diamines. This listingis not intended to be exhaustive of the various classes of couplingagents known in the art but, rather, is exemplary of the more commoncoupling agents. (See Killen and Lindstrom, Jour. Immun. 133:1335-2549(1984); Jansen et al., Immunological Reviews 62:185-216 (1982); andVitetta et al., Science 238:1098 (1987).

In some embodiments, in addition to the compositions and methodsprovided herein, the conjugated activatable antibody can also bemodified for site-specific conjugation through modified amino acidsequences inserted or otherwise included in the activatable antibodysequence. These modified amino acid sequences are designed to allow forcontrolled placement and/or dosage of the conjugated agent within aconjugated activatable antibody. For example, the activatable antibodycan be engineered to include cysteine substitutions at positions onlight and heavy chains that provide reactive thiol groups and do notnegatively impact protein folding and assembly, nor alter antigenbinding. In some embodiments, the activatable antibody can be engineeredto include or otherwise introduce one or more non-natural amino acidresidues within the activatable antibody to provide suitable sites forconjugation. In some embodiments, the activatable antibody can beengineered to include or otherwise introduce enzymatically activatablepeptide sequences within the activatable antibody sequence.

Suitable linkers are described in the literature. (See, for example,Ramakrishnan, S. et al., Cancer Res. 44:201-208 (1984) describing use ofMBS (M-maleimidobenzoyl-N-hydroxysuccinimide ester). See also, U.S. Pat.No. 5,030,719, describing use of halogenated acetyl hydrazide derivativecoupled to an antibody by way of an oligopeptide linker. Particularlysuitable linkers include: (i) SMPT(4-succinimidyloxycarbonyl-alpha-methyl-alpha-(2-pridyl-dithio)-toluene(Pierce Chem. Co., Cat. (21558G); (ii) SPDP (succinimidyl-6[3-(2-pyridyldithio) propionamido]hexanoate (Pierce Chem. Co., Cat#21651G); and (iii) Sulfo-LC-SPDP (sulfosuccinimidyl 6[3-(2-pyridyldithio)-propianamide] hexanoate (Pierce Chem. Co. Cat.#2165-G. Additional linkers include, but are not limited to, SMCC,sulfo-SMCC, SPDB, or sulfo-SPDB.

The linkers described above contain components that have differentattributes, thus leading to conjugates with differing physio-chemicalproperties. For example, the linker SMPT contains a sterically hindereddisulfide bond, and can form conjugates with increased stability.Disulfide linkages, are in general, less stable than other linkagesbecause the disulfide linkage is cleaved in vitro, resulting in lessconjugate available.

The reagent EDC (1-ethyl-3-(3-dimethylaminopropyl)carbodiimidehydrochloride is useful to create a carboxamide starting with acarboxylic acid and a primary or secondary amine. Thus, EDC may be usedto link lysine residues in an antibody with a carboxylic acid in alinker or toxin, or to link aspartate or glutamate residues in anantibody with an amine in a linker or toxin. Such conjugation reactionsutilizing EDC may be enhanced by addition of NHS (N-hydroxysuccinimide)or sulfo-NHS (N-hydroxy-3-oxysulfonylsuccinimide). Addition of NHS orsulfo-NHS to such conjugation reactions may enhance the rate,completeness, selectivity, and/or reproducibility of the conjugationreactions.

In some embodiments, the linkers are cleavable. In some embodiments, thelinkers are non-cleavable. In some embodiments, two or more linkers arepresent. The two or more linkers are all the same, e.g., cleavable ornon-cleavable, or the two or more linkers are different, e.g., at leastone cleavable and at least one non-cleavable.

The present disclosure utilizes several methods for attaching agents toAbs of the anti-CD3ε antibodies, activatable antibodies, multispecificantibodies and/or multispecific activatable antibodies: (a) attachmentto the carbohydrate moieties of the AB, or (b) attachment to sulfhydrylgroups of the AB, or (c) attachment to amino groups of the AB, or (d)attachment to carboxylate groups of the AB. According to the disclosure,ABs may be covalently attached to an agent through an intermediatelinker having at least two reactive groups, one to react with AB and oneto react with the agent. The linker, which may include any compatibleorganic compound, can be chosen such that the reaction with AB (oragent) does not adversely affect AB reactivity and selectivity.Furthermore, the attachment of linker to agent might not destroy theactivity of the agent. Suitable linkers for reaction with oxidizedantibodies or oxidized antibody fragments include those containing anamine selected from the group consisting of primary amine, secondaryamine, hydrazine, hydrazide, hydroxylamine, phenylhydrazine,semicarbazide and thiosemicarbazide groups. Such reactive functionalgroups may exist as part of the structure of the linker, or may beintroduced by suitable chemical modification of linkers not containingsuch groups.

According to the present disclosure, suitable linkers for attachment toreduced ABs of the anti-CD3ε antibodies, activatable antibodies,multispecific antibodies and/or multispecific activatable antibodiesinclude those having certain reactive groups capable of reaction with asulfhydryl group of a reduced antibody or fragment. Such reactive groupsinclude, but are not limited to: reactive haloalkyl groups (including,for example, haloacetyl groups), p-mercuribenzoate groups and groupscapable of Michael-type addition reactions (including, for example,maleimides and groups of the type described by Mitra and Lawton, 1979,J. Amer. Chem. Soc. 101: 3097-3110).

According to the present disclosure, suitable linkers for attachment toneither oxidized nor reduced ABs of the anti-CD3ε antibodies,activatable antibodies, multispecific antibodies and/or multispecificactivatable antibodies include those having certain functional groupscapable of reaction with the primary amino groups present in unmodifiedlysine residues in the AB. Such reactive groups include, but are notlimited to, NHS carboxylic or carbonic esters, sulfo-NHS carboxylic orcarbonic esters, 4-nitrophenyl carboxylic or carbonic esters,pentafluorophenyl carboxylic or carbonic esters, acyl imidazoles,isocyanates, and isothiocyanates.

According to the present disclosure, suitable linkers for attachment toneither oxidized nor reduced ABs include those having certain functionalgroups capable of reaction with the carboxylic acid groups present inaspartate or glutamate residues in the AB, which have been activatedwith suitable reagents. Suitable activating reagents include EDC, withor without added NHS or sulfo-NHS, and other dehydrating agents utilizedfor carboxamide formation. In these instances, the functional groupspresent in the suitable linkers would include primary and secondaryamines, hydrazines, hydroxylamines, and hydrazides.

The agent may be attached to the linker before or after the linker isattached to the AB. In certain applications it may be desirable to firstproduce an AB-linker intermediate in which the linker is free of anassociated agent. Depending upon the particular application, a specificagent may then be covalently attached to the linker. In otherembodiments the AB is first attached to the MM, CM and associatedlinkers and then attached to the linker for conjugation purposes.

Branched Linkers:

In specific embodiments, branched linkers that have multiple sites forattachment of agents are utilized. For multiple site linkers, a singlecovalent attachment to an AB would result in an AB-linker intermediatecapable of binding an agent at a number of sites. The sites may bealdehyde or sulfhydryl groups or any chemical site to which agents canbe attached.

In some embodiments, higher specific activity (or higher ratio of agentsto AB) can be achieved by attachment of a single site linker at aplurality of sites on the AB. This plurality of sites may be introducedinto the AB by either of two methods. First, one may generate multiplealdehyde groups and/or sulfhydryl groups in the same AB. Second, one mayattach to an aldehyde or sulfhydryl of the AB a “branched linker” havingmultiple functional sites for subsequent attachment to linkers. Thefunctional sites of the branched linker or multiple site linker may bealdehyde or sulfhydryl groups, or may be any chemical site to whichlinkers may be attached. Still higher specific activities may beobtained by combining these two approaches; that is, attaching multiplesite linkers at several sites on the AB.

Cleavable Linkers:

Peptide linkers that are susceptible to cleavage by enzymes of thecomplement system, such as but not limited to urokinase, tissueplasminogen activator, trypsin, plasmin, or another enzyme havingproteolytic activity may be used in one embodiment of the presentdisclosure. According to one method of the present disclosure, an agentis attached via a linker susceptible to cleavage by complement. Theantibody is selected from a class that can activate complement. Theantibody-agent conjugate, thus, activates the complement cascade andreleases the agent at the target site. According to another method ofthe present disclosure, an agent is attached via a linker susceptible tocleavage by enzymes having a proteolytic activity such as a urokinase, atissue plasminogen activator, plasmin, or trypsin. These cleavablelinkers are useful in conjugated activatable antibodies that include anextracellular toxin, e.g., by way of non-limiting example, any of theextracellular toxins shown in Table 4.

Non-liming examples of cleavable linker sequences are provided in Table5.

TABLE 5  Exemplary Linker Sequences for ConjugationTypes of Cleavable Sequences Amino Acid SequencePlasmin cleavable sequences Pro-urokinase PRFKIIGG (SEQ ID NO: 342)PRFRIIGG (SEQ ID NO: 343) TGFβ SSRHRRALD (SEQ ID NO: 344) PlasminogenRKSSIIIRMRDVVL (SEQ ID NO: 345) StaphylokinaseSSSFDKGKYKKGDDA (SEQ ID NO: 346) SSSFDKGKYKRGDDA (SEQ ID NO: 347)Factor Xa cleavable sequences IEGR (SEQ ID NO: 348)IDGR (SEQ ID NO: 349) GGSIDGR (SEQ ID NO: 350) MMP cleavable sequencesGelatinase A PLGLWA (SEQ ID NO: 351) Collagenase cleavable sequencesCalf skin collagen (α1(I) chain) GPQGIAGQ (SEQ ID NO: 352)Calf skin collagen (α2(I) chain) GPQGLLGA (SEQ ID NO: 353)Bovine cartilage collagen (α1(II) chain) GIAGQ (SEQ ID NO: 354)Human liver collagen (α1(III) chain) GPLGIAGI (SEQ ID NO: 355) Human α₂MGPEGLRVG (SEQ ID NO: 356) Human PZP YGAGLGVV (SEQ ID NO: 357)AGLGVVER (SEQ ID NO: 358) AGLGISST (SEQ ID NO: 359) Rat α₁MEPQALAMS (SEQ ID NO: 360) QALAMSAI (SEQ ID NO: 361) Rat α₂MAAYHLVSQ (SEQ ID NO: 362) MDAFLESS (SEQ ID NO: 363) Rat α₁I₃(2J)ESLPVVAV (SEQ ID NO: 364) Rat α₁I₃(27J) SAPAVESE (SEQ ID NO: 365)Human fibroblast collagenase DVAQFVLT (SEQ ID NO: 366)(autolytic cleavages) VAQFVLTE (SEQ ID NO: 367)AQFVLTEG (SEQ ID NO: 368) PVQPIGPQ (SEQ ID NO: 369)

In addition, agents may be attached via disulfide bonds (for example,the disulfide bonds on a cysteine molecule) to the AB. Since many tumorsnaturally release high levels of glutathione (a reducing agent) this canreduce the disulfide bonds with subsequent release of the agent at thesite of delivery. In certain specific embodiments the reducing agentthat would modify a CM would also modify the linker of the conjugatedactivatable antibody.

Spacers and Cleavable Elements:

In still another embodiment, it may be necessary to construct the linkerin such a way as to optimize the spacing between the agent and the AB ofthe activatable antibody. This may be accomplished by use of a linker ofthe general structure:

W—(CH₂)n-Q

whereinW is either —NH—CH₂— or —CH₂—;Q is an amino acid, peptide; andn is an integer from 0 to 20.

In still other embodiments, the linker may comprise a spacer element anda cleavable element. The spacer element serves to position the cleavableelement away from the core of the AB such that the cleavable element ismore accessible to the enzyme responsible for cleavage. Certain of thebranched linkers described above may serve as spacer elements.

Throughout this discussion, it should be understood that the attachmentof linker to agent (or of spacer element to cleavable element, orcleavable element to agent) need not be particular mode of attachment orreaction. Any reaction providing a product of suitable stability andbiological compatibility is acceptable.

Serum Complement and Selection of Linkers:

According to one method of the present disclosure, when release of anagent is desired, an AB that is an antibody of a class that can activatecomplement is used. The resulting conjugate retains both the ability tobind antigen and activate the complement cascade. Thus, according tothis embodiment of the present disclosure, an agent is joined to one endof the cleavable linker or cleavable element and the other end of thelinker group is attached to a specific site on the AB. For example, ifthe agent has an hydroxy group or an amino group, it may be attached tothe carboxy terminus of a peptide, amino acid or other suitably chosenlinker via an ester or amide bond, respectively. For example, suchagents may be attached to the linker peptide via a carbodimide reaction.If the agent contains functional groups that would interfere withattachment to the linker, these interfering functional groups can beblocked before attachment and deblocked once the product conjugate orintermediate is made. The opposite or amino terminus of the linker isthen used either directly or after further modification for binding toan AB that is capable of activating complement.

Linkers (or spacer elements of linkers) may be of any desired length,one end of which can be covalently attached to specific sites on the ABof the activatable antibody. The other end of the linker or spacerelement may be attached to an amino acid or peptide linker.

Thus when these conjugates bind to antigen in the presence of complementthe amide or ester bond that attaches the agent to the linker will becleaved, resulting in release of the agent in its active form. Theseconjugates, when administered to a subject, will accomplish delivery andrelease of the agent at the target site, and are particularly effectivefor the in vivo delivery of pharmaceutical agents, antibiotics,antimetabolites, antiproliferative agents and the like as presented inbut not limited to those in Table 4.

Linkers for Release without Complement Activation:

In yet another application of targeted delivery, release of the agentwithout complement activation is desired since activation of thecomplement cascade will ultimately lyse the target cell. Hence, thisapproach is useful when delivery and release of the agent should beaccomplished without killing the target cell. Such is the goal whendelivery of cell mediators such as hormones, enzymes, corticosteroids,neurotransmitters, genes or enzymes to target cells is desired. Theseconjugates may be prepared by attaching the agent to an AB that is notcapable of activating complement via a linker that is mildly susceptibleto cleavage by serum proteases. When this conjugate is administered toan individual, antigen-antibody complexes will form quickly whereascleavage of the agent will occur slowly, thus resulting in release ofthe compound at the target site.

Biochemical Cross Linkers:

In other embodiments, the activatable antibody may be conjugated to oneor more therapeutic agents using certain biochemical cross-linkers.Cross-linking reagents form molecular bridges that tie togetherfunctional groups of two different molecules. To link two differentproteins in a step-wise manner, hetero-bifunctional cross-linkers can beused that eliminate unwanted homopolymer formation.

Peptidyl linkers cleavable by lysosomal proteases are also useful, forexample, Val-Cit, Val-Ala or other dipeptides. In addition, acid-labilelinkers cleavable in the low-pH environment of the lysosome may be used,for example: bis-sialyl ether. Other suitable linkers includecathepsin-labile substrates, particularly those that show optimalfunction at an acidic pH.

Exemplary hetero-bifunctional cross-linkers are referenced in Table 6.

TABLE 6 Exemplary Hetero-Bifunctional Cross Linkers HETERO-BIFUNCTIONALCROSS-LINKERS Spacer Arm Length after cross-linking Linker ReactiveToward Advantages and Applications (Angstroms) SMPT Primary aminesGreater stability 11.2 Å Sulfhydryls SPDP Primary amines Thiolation  6.8Å Sulfhydryls Cleavable cross-linking LC-SPDP Primary amines Extendedspacer arm 15.6 Å Sulfhydryls Sulfo-LC-SPDP Primary amines Extenderspacer arm 15.6 Å Sulfhydryls Water-soluble SMCC Primary amines Stablemaleimide reactive 11.6 Å group Sulfhydryls Enzyme-antibody conjugationHapten-carrier protein conjugation Sulfo-SMCC Primary amines Stablemaleimide reactive 11.6 Å group Sulfhydryls Water-solubleEnzyme-antibody conjugation MBS Primary amines Enzyme-antibodyconjugation  9.9 Å Sulfhydryls Hapten-carrier protein conjugationSulfo-MBS Primary amines Water-soluble  9.9 Å Sulfhydryls SIAB Primaryamines Enzyme-antibody conjugation 10.6 Å Sulfhydryls Sulfo-SIAB Primaryamines Water-soluble 10.6 Å Sulfhydryls SMPB Primary amines Extendedspacer arm 14.5 Å Sulfhydryls Enzyme-antibody conjugation Sulfo-SMPBPrimary amines Extended spacer arm 14.5 Å Sulfhydryls Water-solubleEDE/Sulfo-NHS Primary amines Hapten-Carrier conjugation 0 Carboxylgroups ABH Carbohydrates Reacts with sugar groups 11.9 Å Nonselective

Non-Cleavable Linkers or Direct Attachment:

In still other embodiments of the disclosure, the conjugate may bedesigned so that the agent is delivered to the target but not released.This may be accomplished by attaching an agent to an AB either directlyor via a non-cleavable linker.

These non-cleavable linkers may include amino acids, peptides, D-aminoacids or other organic compounds that may be modified to includefunctional groups that can subsequently be utilized in attachment to ABsby the methods described herein. A-general formula for such an organiclinker could be

W—(CH₂)n-Q

whereinW is either —NH—CH₂— or —CH₂—;Q is an amino acid, peptide; andn is an integer from 0 to 20.

Non-Cleavable Conjugates:

In some embodiments, a compound may be attached to ABs that do notactivate complement. When using ABs that are incapable of complementactivation, this attachment may be accomplished using linkers that aresusceptible to cleavage by activated complement or using linkers thatare not susceptible to cleavage by activated complement.

The antibodies disclosed herein can also be formulated asimmunoliposomes. Liposomes containing the antibody are prepared bymethods known in the art, such as described in Epstein et al., Proc.Natl. Acad. Sci. USA, 82: 3688 (1985); Hwang et al., Proc. Natl Acad.Sci. USA, 77: 4030 (1980); and U.S. Pat. Nos. 4,485,045 and 4,544,545.Liposomes with enhanced circulation time are disclosed in U.S. Pat. No.5,013,556.

Particularly useful liposomes can be generated by the reverse-phaseevaporation method with a lipid composition comprisingphosphatidylcholine, cholesterol, and PEG-derivatizedphosphatidylethanolamine (PEG-PE). Liposomes are extruded throughfilters of defined pore size to yield liposomes with the desireddiameter. Fab′ fragments of the antibody of the present disclosure canbe conjugated to the liposomes as described in Martin et al., J. Biol.Chem., 257: 286-288 (1982) via a disulfide-interchange reaction.

Activatable Antibodies and Multispecific Activatable Antibodies HavingNon-Binding Steric Moieties or Binding Partners for Non-Binding StericMoieties

The disclosure also provides activatable antibodies and/or multispecificactivatable antibodies that include non-binding steric moieties (NB) orbinding partners (BP) for non-binding steric moieties, where the BPrecruits or otherwise attracts the NB to the activatable antibody and/ormultispecific activatable antibody. The activatable antibodies providedherein include, for example, an activatable antibody that includes anon-binding steric moiety (NB), a cleavable linker (CL) and an antibodyor antibody fragment (AB) that binds a first target or epitope; amultispecific activatable antibody that includes a binding partner for anon-binding steric moiety (BP), a CL and an AB; and a multispecificactivatable antibody that includes a BP to which an NB has beenrecruited, a CL and AB that binds a first target or epitope. Themultispecific activatable antibodies provided herein include, forexample, a multispecific activatable antibody that includes anon-binding steric moiety (NB), a cleavable linker (CL) and at least afirst antibody or antibody fragment (AB1) that binds a first target orepitope; a multispecific activatable antibody that includes a bindingpartner for a non-binding steric moiety (BP), a CL and an AB1; and amultispecific activatable antibody that includes a BP to which an NB hasbeen recruited, a CL and AB1 that binds a first target or epitope.Activatable antibodies in which the NB is covalently linked to the CLand AB or is associated by interaction with a BP that is covalentlylinked to the CL and AB are referred to herein as “NB-containingactivatable antibodies.” Multispecific activatable antibodies in whichthe NB is covalently linked to the CL and AB1 or is associated byinteraction with a BP that is covalently linked to the CL and AB1 arereferred to herein as “NB-containing multispecific activatableantibodies.” By activatable or switchable is meant that the activatableantibody exhibits a first level of binding to a target when theactivatable antibody is in an inhibited, masked or uncleaved state(i.e., a first conformation), and a second level of binding to thetarget when the activatable antibody is in an uninhibited, unmaskedand/or cleaved state (i.e., a second conformation, i.e., activatedantibody), where the second level of target binding is greater than thefirst level of target binding. The activatable antibody and/ormultispecific activatable antibody compositions can exhibit increasedbioavailability and more favorable biodistribution compared toconventional antibody therapeutics.

In some embodiments, activatable antibodies and/or multispecificactivatable antibodies provide for reduced toxicity and/or adverse sideeffects that could otherwise result from binding of the activatableantibody and/or multispecific activatable antibody at non-treatmentsites and/or non-diagnostic sites if the activatable antibody and/ormultispecific activatable antibody were not masked or otherwiseinhibited from binding to such a site.

Anti-CD3ε activatable antibodies that include a non-binding stericmoiety (NB) can be made using the methods set forth in PCT PublicationNo. WO 2013/192546, the contents of which are hereby incorporated byreference in their entirety.

Use of Anti-CD3ε, Activatable Antibodies, Multispecific Antibodies andMultispecific Activatable Antibodies

It will be appreciated that administration of therapeutic entities inaccordance with the disclosure will be administered with suitablecarriers, excipients, and other agents that are incorporated intoformulations to provide improved transfer, delivery, tolerance, and thelike. A multitude of appropriate formulations can be found in theformulary known to all pharmaceutical chemists: Remington'sPharmaceutical Sciences (15th ed., Mack Publishing Company, Easton, Pa.(1975)), particularly Chapter 87 by Blaug, Seymour, therein. Theseformulations include, for example, powders, pastes, ointments, jellies,waxes, oils, lipids, lipid (cationic or anionic) containing vesicles(such as Lipofectin™), DNA conjugates, anhydrous absorption pastes,oil-in-water and water-in-oil emulsions, emulsions carbowax(polyethylene glycols of various molecular weights), semi-solid gels,and semi-solid mixtures containing carbowax. Any of the foregoingmixtures may be appropriate in treatments and therapies in accordancewith the present disclosure, provided that the active ingredient in theformulation is not inactivated by the formulation and the formulation isphysiologically compatible and tolerable with the route ofadministration. See also Baldrick P. “Pharmaceutical excipientdevelopment: the need for preclinical guidance.” Regul. ToxicolPharmacol. 32(2):210-8 (2000), Wang W. “Lyophilization and developmentof solid protein pharmaceuticals.” Int. J. Pharm. 203(1-2):1-60 (2000),Charman W N “Lipids, lipophilic drugs, and oral drug delivery-someemerging concepts.” J Pharm Sci. 89(8):967-78 (2000), Powell et al.“Compendium of excipients for parenteral formulations” PDA J Pharm SciTechnol. 52:238-311 (1998) and the citations therein for additionalinformation related to formulations, excipients and carriers well knownto pharmaceutical chemists.

In one embodiment, an anti-CD3ε antibody, an activatable antibody, amultispecific antibody and/or a multispecific activatable antibody ofthe disclosure may be used as therapeutic agents. Such agents willgenerally be employed to diagnose, prognose, monitor, treat, alleviate,and/or prevent a disease or pathology in a subject. A therapeuticregimen is carried out by identifying a subject, e.g., a human patientor other mammal suffering from (or at risk of developing) a disorderusing standard methods. An anti-CD3ε antibody and/or an activatableantibody preparation is administered to the subject and will generallyhave an effect due to its binding with the CD3ε. A multispecificantibody and/or a multispecific activatable antibody preparation, forexample in some embodiments, one having high specificity and highaffinity for its two or more target antigens, is administered to thesubject and will generally have an effect due to its binding with thetargets. Administration of anti-CD3ε antibody, an activatable antibody,a multispecific antibody and/or a multispecific activatable antibody mayactivate T cells via engagement of CD3ε on the T cells. Administrationof anti-CD3ε antibody, an activatable antibody, a multispecific antibodyand/or a multispecific activatable antibody may agonize, stimulate,activate, and/or augment CD3-mediated T cell activation.

Generally, alleviation or treatment of a disease or disorder involvesthe lessening of one or more symptoms or medical problems associatedwith the disease or disorder. For example, in the case of cancer, thetherapeutically effective amount of the drug can accomplish one or acombination of the following: reduce the number of cancer cells; reducethe tumor size; inhibit (i.e., to decrease to some extent and/or stop)cancer cell infiltration into peripheral organs; inhibit tumormetastasis; inhibit, to some extent, tumor growth; and/or relieve tosome extent one or more of the symptoms associated with the cancer. Insome embodiments, a composition of this disclosure can be used toprevent the onset or reoccurrence of the disease or disorder in asubject, e.g., a human or other mammal, such as a non-human primate,companion animal (e.g., cat, dog, horse), farm animal, work animal, orzoo animal. The terms subject and patient are used interchangeablyherein.

A therapeutically effective amount of an anti-CD3ε antibody, anactivatable antibody, a multispecific antibody and/or a multispecificactivatable antibody of the disclosure relates generally to the amountneeded to achieve a therapeutic objective. As noted above, this may be abinding interaction between the anti-CD3ε antibody, the activatableantibody, the multispecific antibody and/or a multispecific activatableantibody and its target antigens that, in certain cases, agonize,stimulate, activate, and/or augment CD3-mediated T cell activation. Theamount required to be administered will furthermore depend on thebinding affinity of the anti-CD3ε antibody, the activatable antibody,the multispecific antibody and/or a multispecific activatable antibodyfor its specific antigen, and will also depend on the rate at which anadministered anti-CD3ε antibody, activatable antibody, multispecificantibody and/or multispecific activatable antibody is depleted from thefree volume other subject to which it is administered. Common ranges fortherapeutically effective dosing of an anti-CD3ε antibody, anactivatable antibody, a multispecific antibody and/or antibody fragmentand/or a multispecific activatable antibody of the disclosure may be, byway of nonlimiting example, from about 0.01 μg/kg body weight to about10 mg/kg body weight. In some embodiments, the therapeutically effectivedosing of an anti-CD3ε antibody, an activatable antibody, amultispecific antibody and/or antibody fragment and/or a multispecificactivatable antibody of the disclosure may be, by way of nonlimitingexample, from about 0.01 mg/kg body weight to about 5-10 mg/kg bodyweight. Common dosing frequencies may range, for example, from twicedaily to once a week.

Efficaciousness of treatment is determined in association with any knownmethod for diagnosing or treating the particular disorder. Methods forthe screening of anti-CD3ε antibodies, activatable antibodies,multispecific antibodies and/or multispecific activatable antibodiesthat possess the desired specificity include, but are not limited to,enzyme linked immunosorbent assay (ELISA) and other immunologicallymediated techniques known within the art.

In another embodiment, an anti-CD3ε antibody, an activatable antibody, amultispecific antibody and/or a multispecific activatable antibodydirected two or more targets are used in methods known within the artrelating to the localization and/or quantitation of the targets (e.g.,for use in measuring levels of one or more of the targets withinappropriate physiological samples, for use in diagnostic methods, foruse in imaging the protein, and the like). In a given embodiment, ananti-CD3ε antibody, an activatable antibody, a multispecific antibodyand/or a multispecific activatable antibody directed two or moretargets, or a derivative, fragment, analog or homolog thereof, thatcontain the antibody derived antigen binding domain, are utilized aspharmacologically active compounds (referred to hereinafter as“Therapeutics”).

In another embodiment, an anti-CD3ε antibody, an activatable antibody, amultispecific antibody and/or a multispecific activatable antibodydirected two or more targets is used to isolate one or more of thetargets by standard techniques, such as immunoaffinity, chromatographyor immunoprecipitation. An anti-CD3ε antibody, an activatable antibody,a multispecific antibody and/or a multispecific activatable antibodydirected two or more targets (or a fragment thereof) are useddiagnostically to monitor protein levels in tissue as part of a clinicaltesting procedure, e.g., to determine the efficacy of a given treatmentregimen. Detection can be facilitated by coupling (i.e., physicallylinking) the antibody to a detectable substance. Examples of detectablesubstances include various enzymes, prosthetic groups, fluorescentmaterials, luminescent materials, bioluminescent materials, andradioactive materials. Examples of suitable enzymes include horseradishperoxidase, alkaline phosphatase, β-galactosidase, oracetylcholinesterase; examples of suitable prosthetic group complexesinclude streptavidin/biotin and avidin/biotin; examples of suitablefluorescent materials include umbelliferone, fluorescein, fluoresceinisothiocyanate, rhodamine, dichlorotriazinylamine fluorescein, dansylchloride or phycoerythrin; an example of a luminescent material includesluminol; examples of bioluminescent materials include luciferase,luciferin, and aequorin, and examples of suitable radioactive materialinclude ¹²⁵I, ¹³¹I, ³⁵S or ³H.

In yet another embodiment, an anti-CD3ε antibody, an activatableantibody, a multispecific antibody and/or a multispecific activatableantibody directed two or more targets can be used as an agent fordetecting the presence of one or more of the targets (or a fragmentthereof) in a sample. In some embodiments, the antibody contains adetectable label. Antibodies are polyclonal, or in some embodiments,monoclonal. An intact antibody, or a fragment thereof (e.g., F_(ab),scFv, or F_((ab)2)) is used. The term “labeled”, with regard to theprobe or antibody, is intended to encompass direct labeling of the probeor antibody by coupling (i.e., physically linking) a detectablesubstance to the probe or antibody, as well as indirect labeling of theprobe or antibody by reactivity with another reagent that is directlylabeled. Examples of indirect labeling include detection of a primaryantibody using a fluorescently-labeled secondary antibody andend-labeling of an antibody with biotin such that it can be detectedwith fluorescently-labeled streptavidin. The term “biological sample” isintended to include tissues, cells and biological fluids isolated from asubject, as well as tissues, cells and fluids present within a subject.Included within the usage of the term “biological sample”, therefore, isblood and a fraction or component of blood including blood serum, bloodplasma, or lymph. That is, the detection method of the disclosure can beused to detect a protein in a biological sample in vitro as well as invivo. For example, in vitro techniques for detection of an analyteprotein include enzyme linked immunosorbent assays (ELISAs), Westernblots, immunoprecipitations, and immunofluorescence. Procedures forconducting immunoassays are described, for example in “ELISA: Theory andPractice: Methods in Molecular Biology”, Vol. 42, J. R. Crowther (Ed.)Human Press, Totowa, N. J., 1995; “Immunoassay”, E. Diamandis and T.Christopoulus, Academic Press, Inc., San Diego, Calif., 1996; and“Practice and Theory of Enzyme Immunoassays”, P. Tijssen, ElsevierScience Publishers, Amsterdam, 1985. Furthermore, in vivo techniques fordetection of an analyte protein include introducing into a subject alabeled anti-analyte protein antibody. For example, the antibody can belabeled with a radioactive marker whose presence and location in asubject can be detected by standard imaging techniques.

The anti-CD3ε antibody, the activatable antibody, the multispecificantibodies and/or multispecific activatable antibodies of the disclosureare also useful in a variety of diagnostic and prophylacticformulations. In one embodiment, an anti-CD3ε antibody, an activatableantibody, a multispecific antibody and/or multispecific activatableantibody is administered to patients that are at risk of developing oneor more of the aforementioned disorders. A patient's or organ'spredisposition to one or more of the disorders can be determined usinggenotypic, serological or biochemical markers.

In another embodiment of the disclosure, an anti-CD3ε antibody, anactivatable antibody, a multispecific antibody and/or multispecificactivatable antibody is administered to human individuals diagnosed witha clinical indication associated with one or more of the aforementioneddisorders. Upon diagnosis, an anti-CD3ε antibody, an activatableantibody, a multispecific antibody and/or multispecific activatableantibody is administered to mitigate or reverse the effects of theclinical indication.

Anti-CD3ε antibodies, activatable antibodies, multispecific antibodiesand/or multispecific activatable antibodies are also useful in thedetection of one or more targets in patient samples and accordingly areuseful as diagnostics. For example, the anti-CD3ε antibodies, theactivatable antibodies, the multispecific antibodies and/ormultispecific activatable antibodies of the disclosure are used in invitro assays, e.g., ELISA, to detect one or more target levels in apatient sample.

In one embodiment, an anti-CD3ε antibody, an activatable antibody, amultispecific antibody and/or multispecific activatable antibody isimmobilized on a solid support (e.g., the well(s) of a microtiterplate). The immobilized antibody and/or activatable antibody serves as acapture antibody for any target(s) that may be present in a test sample.Prior to contacting the immobilized anti-CD3ε antibody, activatableantibody, multispecific antibody and/or immobilized multispecificactivatable antibody with a patient sample, the solid support is rinsedand treated with a blocking agent such as milk protein or albumin toprevent nonspecific adsorption of the analyte.

Subsequently the wells are treated with a test sample suspected ofcontaining the antigen, or with a solution containing a standard amountof the antigen. Such a sample is, e.g., a serum sample from a subjectsuspected of having levels of circulating antigen considered to bediagnostic of a pathology. After rinsing away the test sample orstandard, the solid support is treated with a second antibody that isdetectably labeled. The labeled second antibody serves as a detectingantibody. The level of detectable label is measured, and theconcentration of target antigen(s) in the test sample is determined bycomparison with a standard curve developed from the standard samples.

It will be appreciated that based on the results obtained using theanti-CD3ε antibody, the activatable antibody, the multispecific antibodyand/or multispecific activatable antibody in an in vitro diagnosticassay, it is possible to stage a disease in a subject based onexpression levels of the target antigen(s). For a given disease, samplesof blood are taken from subjects diagnosed as being at various stages inthe progression of the disease, and/or at various points in thetherapeutic treatment of the disease. Using a population of samples thatprovides statistically significant results for each stage of progressionor therapy, a range of concentrations of the antigen that may beconsidered characteristic of each stage is designated.

Anti-CD3ε antibodies, activatable antibodies, multispecific antibodiesand/or multispecific activatable antibodies can also be used indiagnostic and/or imaging methods. In some embodiments, such methods arein vitro methods. In some embodiments, such methods are in vivo methods.In some embodiments, such methods are in situ methods. In someembodiments, such methods are ex vivo methods. For example, activatableantibodies and/or multispecific activatable antibodies having anenzymatically cleavable CM can be used to detect the presence or absenceof an enzyme that is capable of cleaving the CM. Such activatableantibodies and/or multispecific activatable antibodies can be used indiagnostics, which can include in vivo detection (e.g., qualitative orquantitative) of enzyme activity (or, in some embodiments, anenvironment of increased reduction potential such as that which canprovide for reduction of a disulfide bond) through measured accumulationof multispecific activated antibodies (i.e., antibodies resulting fromcleavage of an activatable antibody and/or a multispecific activatableantibody) in a given cell or tissue of a given host organism. Suchaccumulation of activated multispecific antibodies indicates not onlythat the tissue expresses enzymatic activity (or an increased reductionpotential depending on the nature of the CM) but also that the tissueexpresses at least one target to which the activated antibody binds.

For example, the CM can be selected to be a protease substrate for aprotease found at the site of a tumor, at the site of a viral orbacterial infection at a biologically confined site (e.g., such as in anabscess, in an organ, and the like), and the like. At least one of theAB can be one that binds a target antigen. Using methods familiar to oneskilled in the art, a detectable label (e.g., a fluorescent label orradioactive label or radiotracer) can be conjugated to an AB or otherregion of a multispecific antibody and/or multispecific activatableantibody. Suitable detectable labels are discussed in the context of theabove screening methods and additional specific examples are providedbelow. Using at least one AB specific to a protein or peptide of thedisease state, along with a protease whose activity is elevated in thedisease tissue of interest, activatable antibodies will exhibit anincreased rate of binding to disease tissue relative to tissues wherethe CM specific enzyme is not present at a detectable level or ispresent at a lower level than in disease tissue or is inactive (e.g., inzymogen form or in complex with an inhibitor). Since small proteins andpeptides are rapidly cleared from the blood by the renal filtrationsystem, and because the enzyme specific for the CM is not present at adetectable level (or is present at lower levels in non-disease tissuesor is present in inactive conformation), accumulation of activatedmultispecific antibodies in the disease tissue is enhanced relative tonon-disease tissues.

In another example, activatable antibodies and/or activatablemultispecific antibodies can be used to detect the presence or absenceof a cleaving agent in a sample. For example, where the activatableantibodies and/or multispecific activatable antibodies contain a CMsusceptible to cleavage by an enzyme, the multispecific activatableantibodies can be used to detect (either qualitatively orquantitatively) the presence of an enzyme in the sample. In anotherexample, where the activatable antibodies and/or multispecificactivatable antibodies contain a CM susceptible to cleavage by reducingagent, the activatable antibodies and/or multispecific activatableantibodies can be used to detect (either qualitatively orquantitatively) the presence of reducing conditions in a sample. Tofacilitate analysis in these methods, the activatable antibodies and/ormultispecific activatable antibodies can be detectably labeled, and canbe bound to a support (e.g., a solid support, such as a slide or bead).The detectable label can be positioned on a portion of the activatableantibody and/or multispecific activatable antibody that is not releasedfollowing cleavage, for example, the detectable label can be a quenchedfluorescent label or other label that is not detectable until cleavagehas occurred. The assay can be conducted by, for example, contacting theimmobilized, detectably labeled activatable antibodies and/ormultispecific activatable antibodies with a sample suspected ofcontaining an enzyme and/or reducing agent for a time sufficient forcleavage to occur, then washing to remove excess sample andcontaminants. The presence or absence of the cleaving agent (e.g.,enzyme or reducing agent) in the sample is then assessed by a change indetectable signal of the activatable antibodies and/or multispecificactivatable antibodies prior to contacting with the sample e.g., thepresence of and/or an increase in detectable signal due to cleavage ofthe activatable antibody and/or multispecific activatable antibody bythe cleaving agent in the sample.

Such detection methods can be adapted to also provide for detection ofthe presence or absence of a target that is capable of binding at leastone AB of the multispecific activatable antibodies when cleaved. Thus,the assays can be adapted to assess the presence or absence of acleaving agent and the presence or absence of a target of interest. Thepresence or absence of the cleaving agent can be detected by thepresence of and/or an increase in detectable label of the multispecificactivatable antibodies as described above, and the presence or absenceof the target can be detected by detection of a target-AB complex e.g.,by use of a detectably labeled anti-target antibody.

Activatable antibodies and/or multispecific activatable antibodies arealso useful in in situ imaging for the validation of activatableantibody activation, e.g., by protease cleavage, and binding to aparticular target. In situ imaging is a technique that enableslocalization of proteolytic activity and target in biological samplessuch as cell cultures or tissue sections. Using this technique, it ispossible to confirm both binding to a given target and proteolyticactivity based on the presence of a detectable label (e.g., afluorescent label).

These techniques are useful with any frozen cells or tissue derived froma disease site (e.g. tumor tissue) or healthy tissues. These techniquesare also useful with fresh cell or tissue samples.

In these techniques, an activatable antibody and/or multispecificactivatable antibody is labeled with a detectable label. The detectablelabel may be a fluorescent dye, (e.g. a fluorophore, FluoresceinIsothiocyanate (FITC), Rhodamine Isothiocyanate (TRITC), an Alexa Fluor®label), a near infrared (NIR) dye (e.g., Qdot® nanocrystals), acolloidal metal, a hapten, a radioactive marker, biotin and anamplification reagent such as streptavidin, or an enzyme (e.g.horseradish peroxidase or alkaline phosphatase).

Detection of the label in a sample that has been incubated with thelabeled, multispecific activatable antibody indicates that the samplecontains the target and contains a protease that is specific for the CMof the activatable antibody and/or multispecific activatable antibody.In some embodiments, the presence of the protease can be confirmed usingbroad spectrum protease inhibitors such as those described herein,and/or by using an agent that is specific for the protease, for example,an antibody such as A11, which is specific for the protease matriptase(MT-SP1) and inhibits the proteolytic activity of matriptase; see e.g.,International Publication Number WO 2010/129609, published 11 Nov. 2010.The same approach of using broad spectrum protease inhibitors such asthose described herein, and/or by using a more selective inhibitoryagent can be used to identify a protease or class of proteases specificfor the CM of the activatable antibody and/or multispecific activatableantibody. In some embodiments, the presence of the target can beconfirmed using an agent that is specific for the target or thedetectable label can be competed with unlabeled target. In someembodiments, unlabeled activatable antibody and/or multispecificactivatable antibody could be used, with detection by a labeledsecondary antibody or more complex detection system.

Similar techniques are also useful for in vivo imaging where detectionof the fluorescent signal in a subject, e.g., a mammal, including ahuman, indicates that the disease site contains the target and containsa protease that is specific for the CM of the activatable antibodyand/or multispecific activatable antibody.

These techniques are also useful in kits and/or as reagents for thedetection, identification or characterization of protease activity in avariety of cells, tissues, and organisms based on the protease-specificCM in the multispecific activatable antibody.

Therapeutic Administration and Formulations of Anti-CD3ε Antibodies,Activatable Antibodies, Multispecific Antibodies and/or MultispecificActivatable Antibodies

It will be appreciated that administration of therapeutic entities inaccordance with the disclosure will be administered with suitablecarriers, excipients, and other agents that are incorporated intoformulations to provide improved transfer, delivery, tolerance, and thelike. A multitude of appropriate formulations can be found in theformulary known to all pharmaceutical chemists: Remington'sPharmaceutical Sciences (15th ed., Mack Publishing Company, Easton, Pa.(1975)), particularly Chapter 87 by Blaug, Seymour, therein. Theseformulations include, for example, powders, pastes, ointments, jellies,waxes, oils, lipids, lipid (cationic or anionic) containing vesicles(such as Lipofectin™), DNA conjugates, anhydrous absorption pastes,oil-in-water and water-in-oil emulsions, emulsions carbowax(polyethylene glycols of various molecular weights), semi-solid gels,and semi-solid mixtures containing carbowax. Any of the foregoingmixtures may be appropriate in treatments and therapies in accordancewith the present disclosure, provided that the active ingredient in theformulation is not inactivated by the formulation and the formulation isphysiologically compatible and tolerable with the route ofadministration. See also Baldrick P. “Pharmaceutical excipientdevelopment: the need for preclinical guidance.” Regul. ToxicolPharmacol. 32(2):210-8 (2000), Wang W. “Lyophilization and developmentof solid protein pharmaceuticals.” Int. J. Pharm. 203(1-2):1-60 (2000),Charman W N “Lipids, lipophilic drugs, and oral drug delivery-someemerging concepts.” J Pharm Sci. 89(8):967-78 (2000), Powell et al.“Compendium of excipients for parenteral formulations” PDA J Pharm SciTechnol. 52:238-311 (1998) and the citations therein for additionalinformation related to formulations, excipients and carriers well knownto pharmaceutical chemists.

In some embodiments, the anti-CD3ε antibodies, the conjugated anti-CD3εantibodies, the activatable antibodies, the conjugated activatableantibodies, the multispecific antibodies, the multispecific activatableantibodies and/or the conjugated multispecific activatable antibodycompositions—referred to collectively herein as the Therapeutic(s)—areadministered in conjunction with one or more additional agents, or acombination of additional agents. Suitable additional agents includecurrent pharmaceutical and/or surgical therapies for an intendedapplication. For example, the Therapeutic(s) can be used in conjunctionwith an additional chemotherapeutic or anti-neoplastic agent. Forexample, the Therapeutic(s) and additional agent are formulated into asingle therapeutic composition, and the Therapeutic(s) and additionalagent are administered simultaneously. In some embodiments, theTherapeutic(s) and additional agent are separate from each other, e.g.,each is formulated into a separate therapeutic composition, and theTherapeutic(s) and the additional agent are administered simultaneously,or the Therapeutic(s) and the additional agent are administered atdifferent times during a treatment regimen. For example, theTherapeutic(s) is administered prior to the administration of theadditional agent, the Therapeutic(s) is administered subsequent to theadministration of the additional agent, or the Therapeutic(s) and theadditional agent are administered in an alternating fashion. Asdescribed herein, the Therapeutic(s) and additional agent areadministered in single doses or in multiple doses.

In some embodiments, the additional agent is coupled or otherwiseattached to the Therapeutic(s).

Suitable additional agents are selected according to the purpose of theintended application (i.e., killing, prevention of cell proliferation,hormone therapy or gene therapy). Such agents may include but is notlimited to, for example, pharmaceutical agents, toxins, fragments oftoxins, alkylating agents, enzymes, antibiotics, antimetabolites,antiproliferative agents, hormones, neurotransmitters, DNA, RNA, siRNA,oligonucleotides, antisense RNA, aptamers, diagnostics, radiopaque dyes,radioactive isotopes, fluorogenic compounds, magnetic labels,nanoparticles, marker compounds, lectins, compounds that alter cellmembrane permeability, photochemical compounds, small molecules,liposomes, micelles, gene therapy vectors, viral vectors, and the like.Finally, combinations of agents or combinations of different classes ofagents may be used.

The anti-CD3ε antibodies, the conjugated anti-CD3ε antibodies, theactivatable antibodies, the conjugated activatable antibodies, themultispecific antibodies, the multispecific activatable antibodiesand/or the conjugated multispecific activatable antibody compositions ofthe disclosure (also referred to herein as “Therapeutic(s)” or “activecompounds”), and derivatives, fragments, analogs and homologs thereof,can be incorporated into pharmaceutical compositions suitable foradministration. Principles and considerations involved in preparing suchcompositions, as well as guidance in the choice of components areprovided, for example, in Remington's Pharmaceutical Sciences: TheScience And Practice Of Pharmacy 19th ed. (Alfonso R. Gennaro, et al.,editors) Mack Pub. Co., Easton, Pa.: 1995; Drug Absorption Enhancement:Concepts, Possibilities, Limitations, And Trends, Harwood AcademicPublishers, Langhorne, Pa., 1994; and Peptide And Protein Drug Delivery(Advances In Parenteral Sciences, Vol. 4), 1991, M. Dekker, New York.

Such compositions typically comprise the anti-CD3ε antibodies, theconjugated anti-CD3ε antibodies, the activatable antibodies, theconjugated activatable antibodies, the multispecific antibodies, themultispecific activatable antibodies and/or the conjugated multispecificactivatable antibody compositions and a pharmaceutically acceptablecarrier. Where an antibody, activatable antibody, multispecific antibodyand/or a multispecific activatable antibody includes a fragment of theAB domain, the smallest fragment of the AB that specifically binds tothe binding domain of the target protein can be used. For example, basedupon the variable-region sequences of an antibody, peptide molecules canbe designed that retain the ability of the AB to bind the target proteinsequence. Such peptides can be synthesized chemically and/or produced byrecombinant DNA technology. (See, e.g., Marasco et al., Proc. Natl.Acad. Sci. USA, 90: 7889-7893 (1993)).

As used herein, the term “pharmaceutically acceptable carrier” isintended to include any and all solvents, dispersion media, coatings,antibacterial and antifungal agents, isotonic and absorption delayingagents, and the like, compatible with pharmaceutical administration.Suitable carriers are described in the most recent edition ofRemington's Pharmaceutical Sciences, a standard reference text in thefield, which is incorporated herein by reference. Suitable examples ofsuch carriers or diluents include, but are not limited to, water,saline, ringer's solutions, dextrose solution, and 5% human serumalbumin. Liposomes and non-aqueous vehicles such as fixed oils may alsobe used. The use of such media and agents for pharmaceutically activesubstances is well known in the art. Except insofar as any conventionalmedia or agent is incompatible with the active compound, use thereof inthe compositions is contemplated.

The formulations to be used for in vivo administration must be sterile.This is readily accomplished by filtration through sterile filtrationmembranes.

A pharmaceutical composition of the disclosure is formulated to becompatible with its intended route of administration. Examples of routesof administration include parenteral, e.g., intravenous, intradermal,subcutaneous, oral (e.g., inhalation), transdermal (i.e., topical),transmucosal, and rectal administration. Solutions or suspensions usedfor parenteral, intradermal, or subcutaneous application can include thefollowing components: a sterile diluent such as water for injection,saline solution, fixed oils, polyethylene glycols, glycerine, propyleneglycol or other synthetic solvents; antibacterial agents such as benzylalcohol or methyl parabens; antioxidants such as ascorbic acid or sodiumbisulfite; chelating agents such as ethylenediaminetetraacetic acid(EDTA); buffers such as acetates, citrates or phosphates, and agents forthe adjustment of tonicity such as sodium chloride or dextrose. The pHcan be adjusted with acids or bases, such as hydrochloric acid or sodiumhydroxide. The parenteral preparation can be enclosed in ampoules,disposable syringes or multiple dose vials made of glass or plastic.

Pharmaceutical compositions suitable for injectable use include sterileaqueous solutions (where water soluble) or dispersions and sterilepowders for the extemporaneous preparation of sterile injectablesolutions or dispersion. For intravenous administration, suitablecarriers include physiological saline, bacteriostatic water, CremophorEL™ (BASF, Parsippany, N.J.) or phosphate buffered saline (PBS). In allcases, the composition must be sterile and should be fluid to the extentthat easy syringeability exists. It must be stable under the conditionsof manufacture and storage and must be preserved against thecontaminating action of microorganisms such as bacteria and fungi. Thecarrier can be a solvent or dispersion medium containing, for example,water, ethanol, polyol (for example, glycerol, propylene glycol, andliquid polyethylene glycol, and the like), and suitable mixturesthereof. The proper fluidity can be maintained, for example, by the useof a coating such as lecithin, by the maintenance of the requiredparticle size in the case of dispersion and by the use of surfactants.Prevention of the action of microorganisms can be achieved by variousantibacterial and antifungal agents, for example, parabens,chlorobutanol, phenol, ascorbic acid, thimerosal, and the like. In manycases, it will be suitable to include isotonic agents, for example,sugars, polyalcohols such as manitol, sorbitol, sodium chloride in thecomposition. Prolonged absorption of the injectable compositions can bebrought about by including in the composition an agent that delaysabsorption, for example, aluminum monostearate and gelatin.

Sterile injectable solutions can be prepared by incorporating the activecompound in the required amount in an appropriate solvent with one or acombination of ingredients enumerated above, as required, followed byfiltered sterilization. Generally, dispersions are prepared byincorporating the active compound into a sterile vehicle that contains abasic dispersion medium and the required other ingredients from thoseenumerated above. In the case of sterile powders for the preparation ofsterile injectable solutions, methods of preparation are vacuum dryingand freeze-drying that yields a powder of the active ingredient plus anyadditional desired ingredient from a previously sterile-filteredsolution thereof.

Oral compositions generally include an inert diluent or an ediblecarrier. They can be enclosed in gelatin capsules or compressed intotablets. For the purpose of oral therapeutic administration, the activecompound can be incorporated with excipients and used in the form oftablets, troches, or capsules. Oral compositions can also be preparedusing a fluid carrier for use as a mouthwash, wherein the compound inthe fluid carrier is applied orally and swished and expectorated orswallowed. Pharmaceutically compatible binding agents, and/or adjuvantmaterials can be included as part of the composition. The tablets,pills, capsules, troches and the like can contain any of the followingingredients, or compounds of a similar nature: a binder such asmicrocrystalline cellulose, gum tragacanth or gelatin; an excipient suchas starch or lactose, a disintegrating agent such as alginic acid,Primogel, or corn starch; a lubricant such as magnesium stearate orSterotes; a glidant such as colloidal silicon dioxide; a sweeteningagent such as sucrose or saccharin; or a flavoring agent such aspeppermint, methyl salicylate, or orange flavoring.

For administration by inhalation, the compounds are delivered in theform of an aerosol spray from pressured container or dispenser thatcontains a suitable propellant, e.g., a gas such as carbon dioxide, or anebulizer.

Systemic administration can also be by transmucosal or transdermalmeans. For transmucosal or transdermal administration, penetrantsappropriate to the barrier to be permeated are used in the formulation.Such penetrants are generally known in the art, and include, forexample, for transmucosal administration, detergents, bile salts, andfusidic acid derivatives. Transmucosal administration can beaccomplished through the use of nasal sprays or suppositories. Fortransdermal administration, the active compounds are formulated intoointments, salves, gels, or creams as generally known in the art.

The compounds can also be prepared in the form of suppositories (e.g.,with conventional suppository bases such as cocoa butter and otherglycerides) or retention enemas for rectal delivery.

In one embodiment, the active compounds are prepared with carriers thatwill protect the compound against rapid elimination from the body, suchas sustained/controlled release formulations, including implants andmicroencapsulated delivery systems. Biodegradable, biocompatiblepolymers can be used, such as ethylene vinyl acetate, polyanhydrides,polyglycolic acid, collagen, polyorthoesters, and polylactic acid.Methods for preparation of such formulations will be apparent to thoseskilled in the art.

For example, the active ingredients can be entrapped in microcapsulesprepared, for example, by coacervation techniques or by interfacialpolymerization, for example, hydroxymethylcellulose orgelatin-microcapsules and poly-(methylmethacrylate) microcapsules,respectively, in colloidal drug delivery systems (for example,liposomes, albumin microspheres, microemulsions, nano-particles, andnanocapsules) or in macroemulsions.

Sustained-release preparations can be prepared. Suitable examples ofsustained-release preparations include semipermeable matrices of solidhydrophobic polymers containing the antibody, which matrices are in theform of shaped articles, e.g., films, or microcapsules. Examples ofsustained-release matrices include polyesters, hydrogels (for example,poly(2-hydroxyethyl-methacrylate), or poly(vinylalcohol)), polylactides(U.S. Pat. No. 3,773,919), copolymers of L-glutamic acid and γethyl-L-glutamate, non-degradable ethylene-vinyl acetate, degradablelactic acid-glycolic acid copolymers such as the LUPRON DEPOT™(injectable microspheres composed of lactic acid-glycolic acid copolymerand leuprolide acetate), and poly-D-(−)-3-hydroxybutyric acid. Whilepolymers such as ethylene-vinyl acetate and lactic acid-glycolic acidenable release of molecules for over 100 days, certain hydrogels releaseproteins for shorter time periods.

The materials can also be obtained commercially from Alza Corporationand Nova Pharmaceuticals, Inc. Liposomal suspensions (includingliposomes targeted to infected cells with monoclonal antibodies to viralantigens) and can also be used as pharmaceutically acceptable carriers.These can be prepared according to methods known to those skilled in theart, for example, as described in U.S. Pat. No. 4,522,811.

It is especially advantageous to formulate oral or parenteralcompositions in dosage unit form for ease of administration anduniformity of dosage. Dosage unit form as used herein refers tophysically discrete units suited as unitary dosages for the subject tobe treated; each unit containing a predetermined quantity of activecompound calculated to produce the desired therapeutic effect inassociation with the required pharmaceutical carrier. The specificationfor the dosage unit forms of the disclosure are dictated by and directlydependent on the unique characteristics of the active compound and theparticular therapeutic effect to be achieved, and the limitationsinherent in the art of compounding such an active compound for thetreatment of individuals.

The pharmaceutical compositions can be included in a container, pack, ordispenser together with instructions for administration.

The formulation can also contain more than one active compound asnecessary for the particular indication being treated, for example,those with complementary activities that do not adversely affect eachother. In some embodiments, or in addition, the composition can comprisean agent that enhances its function, such as, for example, a cytotoxicagent, cytokine, chemotherapeutic agent, or growth-inhibitory agent.Such molecules are suitably present in combination in amounts that areeffective for the purpose intended.

In one embodiment, the active compounds are administered in combinationtherapy, i.e., combined with other agents, e.g., therapeutic agents,that are useful for treating pathological conditions or disorders, suchas autoimmune disorders and inflammatory diseases. The term “incombination” in this context means that the agents are givensubstantially contemporaneously, either simultaneously or sequentially.If given sequentially, at the onset of administration of the secondcompound, the first of the two compounds is still detectable ateffective concentrations at the site of treatment.

For example, the combination therapy can include one or more antibodiesof the disclosure coformulated with, and/or coadministered with, one ormore additional therapeutic agents, e.g., one or more cytokine andgrowth factor inhibitors, immunosuppressants, anti-inflammatory agents,metabolic inhibitors, enzyme inhibitors, and/or cytotoxic or cytostaticagents, as described in more detail below. Furthermore, one or moreantibodies described herein may be used in combination with two or moreof the therapeutic agents described herein. Such combination therapiesmay advantageously utilize lower dosages of the administered therapeuticagents, thus avoiding possible toxicities or complications associatedwith the various monotherapies.

In other embodiments, one or more antibodies of the disclosure can becoformulated with, and/or coadministered with, one or moreanti-inflammatory drugs, immunosuppressants, or metabolic or enzymaticinhibitors. Nonlimiting examples of the drugs or inhibitors that can beused in combination with the antibodies described herein, include, butare not limited to, one or more of: nonsteroidal anti-inflammatorydrug(s) (NSAIDs), e.g., ibuprofen, tenidap, naproxen, meloxicam,piroxicam, diclofenac, and indomethacin; sulfasalazine; corticosteroidssuch as prednisolone; cytokine suppressive anti-inflammatory drug(s)(CSAIDs); inhibitors of nucleotide biosynthesis, e.g., inhibitors ofpurine biosynthesis, folate antagonists (e.g., methotrexate(N-[4-[[(2,4-diamino-6-pteridinyl)methyl] methylamino]benzoyl]-L-glutamic acid); and inhibitors of pyrimidine biosynthesis,e.g., dihydroorotate dehydrogenase (DHODH) inhibitors. Suitabletherapeutic agents for use in combination with the antibodies of thedisclosure include NSAIDs, CSAIDs, (DHODH) inhibitors (e.g.,leflunomide), and folate antagonists (e.g., methotrexate).

Examples of additional inhibitors include one or more of:corticosteroids (oral, inhaled and local injection); immunosuppressants,e.g., cyclosporin, tacrolimus (FK-506); and mTOR inhibitors, e.g.,sirolimus (rapamycin—RAPAMUNE™ or rapamycin derivatives, e.g., solublerapamycin derivatives (e.g., ester rapamycin derivatives, e.g.,CCI-779); agents that interfere with signaling by proinflammatorycytokines such as TNFα or IL-1 (e.g. IRAK, NIK, IKK, p38 or MAP kinaseinhibitors); COX2 inhibitors, e.g., celecoxib, rofecoxib, and variantsthereof; phosphodiesterase inhibitors, e.g., R973401 (phosphodiesteraseType IV inhibitor); phospholipase inhibitors, e.g., inhibitors ofcytosolic phospholipase 2 (cPLA2) (e.g., trifluoromethyl ketoneanalogs); inhibitors of vascular endothelial cell growth factor orgrowth factor receptor, e.g., VEGF inhibitor and/or VEGF-R inhibitor;and inhibitors of angiogenesis. Suitable therapeutic agents for use incombination with the antibodies of the disclosure areimmunosuppressants, e.g., cyclosporin, tacrolimus (FK-506); mTORinhibitors, e.g., sirolimus (rapamycin) or rapamycin derivatives, e.g.,soluble rapamycin derivatives (e.g., ester rapamycin derivatives, e.g.,CCI-779); COX2 inhibitors, e.g., celecoxib and variants thereof; andphospholipase inhibitors, e.g., inhibitors of cytosolic phospholipase 2(cPLA2), e.g., trifluoromethyl ketone analogs.

Additional examples of therapeutic agents that can be combined with anantibody of the disclosure include one or more of: 6-mercaptopurines(6-MP); azathioprine sulphasalazine; mesalazine; olsalazine;chloroquine/hydroxychloroquine (PLAQUENIL®); pencillamine;aurothiornalate (intramuscular and oral); azathioprine; coichicine;beta-2 adrenoreceptor agonists (salbutamol, terbutaline, salmeteral);xanthines (theophylline, arninophylline); cromoglycate; nedocromil;ketotifen; ipratropium and oxitropium; mycophenolate mofetil; adenosineagonists; antithrombotic agents; complement inhibitors; and adrenergicagents.

All publications and patent documents cited herein are incorporatedherein by reference as if each such publication or document wasspecifically and individually indicated to be incorporated herein byreference. Citation of publications and patent documents is not intendedas an admission that any is pertinent prior art, nor does it constituteany admission as to the contents or date of the same. The inventionhaving now been described by way of written description, those of skillin the art will recognize that the invention can be practiced in avariety of embodiments and that the foregoing description and examplesbelow are for purposes of illustration and not limitation of the claimsthat follow.

EXAMPLES Example 1 Activatable Anti-CD3ε Antibody Masking Moieties

This Example describes identification of masking moieties (MM) to reducebinding of activatable anti-CD3ε antibodies to their target.

MACS1 and MACS2 Selections

Anti-CD3ε antibody SP34 (also referred to herein as SP34 antibody) wasused to screen a peptide library inserted into a circularly permutedouter membrane protein OmpX in E. coli with 6×10¹⁰ members, using amethod similar to the bacterial display technology described in PCTInternational Publication Numbers WO 2005/047461, published May 26,2005, and WO 2009/014726, published Jan. 29, 2009. The selectioninvolved two rounds of magnetic activated cell separation (MACS)followed by two different fluorescent activated cell sorting (FACS)based selection strategies. FIG. 1 shows the generic naming scheme forMACS and FACS populations during a selection process: Magnetic activatedcell separations (MACS) are denoted with an “M” and fluorescentactivated cell sorting (FACS) selections are denoted with an “F”. Theorder of the abbreviation indicates the order of selections/sorts andthe number indicates the number of rounds of that type of selection/sortto which that the population has been subjected. For example, an M2F3population has gone through 2 rounds of MACS selections followed by 3rounds of FACS selections. The starting population that undergoesselection to yield a given population is assumed to be N−1 of the rightmost sort indicator in the nomenclature. For example, the populationthat was sorted to produce the M2F3 population is assumed to have beenthe M2F2 population.

The initial MACS (M1) was done using protein-G Dynabeads (InvitrogenCat#: 10003D) and the SP34 antibody (available from BD BiosciencesCat#556610) at a concentration of 100 nM. For the M1, 6×10¹¹ cells werescreened for binding and 6.6×10⁷ cells were collected. The second roundof MACS sorting (M2) was done in a manner similar to the first roundwith the following differences. The SP34 antibody was added to a finalconcentration of 50 nM, and a total of 4.5×10¹¹ cells were screened forbinding and 3.1×10⁵ cells were collected.

FACS Selections Selection #1

For all FACS selections, 50 μl of induced culture were spun down (5 min@ 3000×g) and labeled with the indicated concentration of antibody. Forthe M2, MACS2FACS1 (M2F1), and MACS2FACS2 (M2F2) selections cells werelabeled with 1 nM of SP34-dylight-488 (ThermoScientific Cat#: 53025) in500 μl PBS, 0.5% BSA, and the top 0.5% of positive cells were sorted byfluorescent activated cell sorting (FACS) using a FACSARIAI instrument(BD Biosciences). For the M2 sort, 3×10⁶ cells were sorted and 5000cells were collected. For subsequent sorts, at least 1×10⁶ cells weresorted. The MACS2FACS3 (M2F3) population was sorted as described herein,but with the addition of 100 nM unlabeled isotype control antibody (BDBiosciences Cat#: 556657). The final MACS2FACS4 (M2F4) round of FACSselection was sorted as before but was labeled using unlabeled SP34antibody labeled with an anti-mouse IgG-488 secondary 1:100 (JacksonImmunoResearch Cat#: 315-486-045), resulting in the MACS2FACS5 (M2F5)population. The final M2F5 population was verified to specifically bindSP34 by demonstrating inhibition of 1 nM SP34-dylight-488 binding in thepresence of 100 nM soluble human CD3ε protein (Sino Biologics Cat#:10977-H08H). One hundred individual clones were chosen for sequenceanalysis, and 91 gave interpretable data. The results are summarized inTable 7.

TABLE 7  Selection #1 M2F5 sequences Peptide namePeptide sequence (occurrence) 14995 YSLWGCEWGCDRGLY(73) (SEQ ID NO: 371)15003 GYRWGCEWNCGGITT(8) (SEQ ID NO: 372) 15253YSACEMFGEVECCFC(7) (SEQ ID NO: 373) 15263WYSGGCEAFCGILSS(2) (SEQ ID NO: 374) 15286GYSGGCEFRCYQLYS(1) (SEQ ID NO: 375)

Selection #2

For all FACS selections, 50 μl of induced culture were spun down (5 min@ 3000×g) and labeled with the indicated concentration of antibody. TheM2, M2F1, and M2F2 sorts, cells were labeled with 10 nM SP34-dylight-488in 500 μL PBS, 0.5% BSA, 100 nM unlabeled isotype control antibody asdescribed herein, and the top 10%, 38%, and 60% of binders were sorted,respectively. In each case, approximately 3×10⁶ cells were sorted.Twenty individual clones were chosen for sequence analysis, and 19 gaveinterpretable data. The results are summarized in Table 8.

TABLE 8  Selection # 2 M2F2 sequences Sequence name Peptide Sequence15855 KFCHCGYYCRVCTLK(1) (SEQ ID NO: 376) 15856LGCNNLWGNEFCHPV(1) (SEQ ID NO: 377) 15858GHPCWGNESYCHTHS(1) (SEQ ID NO: 378) 15859GNNKWCNKPCKCWNK(1) (SEQ ID NO: 379) 15860VYYCGGNESLCGERR(1) (SEQ ID NO: 380) 15861WMTFGCEFSCGTDEW(1) (SEQ ID NO: 381) 15862VYHWGCEYDCFFNDM(1) (SEQ ID NO: 382) 15863FSGCCRGWYNCCHRG(1) (SEQ ID NO: 383) 15864FMCQQRMWGNEFCHQ(1) (SEQ ID NO: 384) 15865MMYCGGNEVLCGPRV(1) (SEQ ID NO: 385) 15866TYSKCRYTVKCTKHN(1) (SEQ ID NO: 386) 15867WYSGGCEAFCGILSS(2) (SEQ ID NO: 387) 15868KYHCTRTSACCTKHH(1) (SEQ ID NO: 388) 15869NCFDPYMLLTYSCNS(1) (SEQ ID NO: 389) 15870GMCGNLWGDESRCWW(1) (SEQ ID NO: 390) 15857SCWDPYMMMNYICNI(3) (SEQ ID NO: 391)

On Cell Peptide Characterization

Eight peptides, 5 from the selection #1 M2F5 and 3 from the selection #2M2F2 populations, were selected for further characterization on cell.The individual clones were labeled with 100, 10, 1, and 0.1 nMSP34-dylight-488 in the presence of 100 nM unlabeled isotype controlantibody as described herein. Each clone was separately labeled withyPet-Mona to quantify the cell surface expression level of the peptides,allowing the calculation of expression normalized binding.

${{Expression}\mspace{14mu} {normalized}\mspace{14mu} {binding}} - \frac{{Mean}\mspace{14mu} {Fab}\mspace{14mu} {fluorescence}}{{Mean}\mspace{14mu} {YpetMona}\mspace{14mu} {fluorescence}}$

And it's assumed that:

MeanYpetMona fluorescence=Peptide expression level

To verify the ability of the binding peptides to inhibit SP34'sinteraction with human CD3ε, each clone was labeled with 1 nMSP34-dylight-488 in the presence of 100 nM human CD3ε. The results aresummarized in Table 9. All of the peptides evaluated specifically boundto SP34-dylight-488 and were inhibited from binding by excess free humanCD3ε.

TABLE 9 Expression normalized binding of individual SP34 binding clonesLabel 14995 15003 15253 15263 15286 15860 15864 15865 100 nM 0.51 0.692.53 2.28 1.72 4.55 1.88 2.24 SP34  10 nM 0.11 0.39 0.98 0.69 0.66 1.720.63 1.08 SP34  1 nM 0.13 0.20 0.68 0.35 0.42 0.52 0.36 0.79 SP34  0.1nM 0.08 0.11 0.55 0.24 0.28 0.48 0.15 0.21 SP34  1 nM 0.08 0.12 0.530.34 0.34 0.51 0.30 0.26   SP34 + 100 nM hCD3ε

Example 2 Activatable Anti-CD3ε Antibodies

This Example describes the production of activatable anti-CD3εantibodies of the disclosure.

The expression vector for the SP34scFv(LvHv)-Fc fusion construct wasassembled using standard molecular biology techniques. Briefly, a DNAfragment encoding the SP34 scFv (LvHv) region was amplified with primersCX2005 and CX2008 using a synthesized SP34scFv(LvHv) sequence astemplate. The DNA encoding the Fc domain was amplified from thepFIL-CHIg-hGl vector (Invitrogen) using primers CX2007 and CX2006. Theoverlapping fragments were combined and amplified with primers CX2005and CX2006 and subsequently cloned into the pOP Hyg expression vector (amodified pCDNA3.1 expression vector (Invitrogen)) using the EcoRI andNotI restriction sites to form antibody SP34scFv(LvHv)-Fc. A DNAfragment encoding antibody SP34scFv(HvLv)-Fc was produced in a similarmanner with the following differences. The DNA fragment encoding thePSP34scFv(HvLv) was amplified using the primers CX2001 and CX200,4 andthe DNA encoding the Fc domain was amplified using the primers CX2002and CX2003. The overlapping fragments were combined and amplified usingprimers CX2001 and C2002. All primer sequences are shown in Table 10.Antibodies SP34scFv(LvHv)-Fc and SP34scFv(HvLv)-Fc were found to bind toJurkat T cells with affinities similar to the ability of SP34 IgG tobind to Jurkat T cells. FIGS. 4A-4C depicts scFv(LvHv)-Fc andscFv(HvLv)-Fc antibodies and activatable antibodies of the disclosure.

Antibodies SP34scFv(LvHv)-Fc and SP34scFv(HvLv)-Fc were also tested fortheir ability to bind to cynomolgus macaque (also referred to herein ascyno) CD3ε. To determine if antibodies SP34scFv(LvHv)-Fc andSP34scFv(HvLv)-Fc could bind to cynomolgus CD3ε, an ELISA-based bindingassay was performed. Recombinant cynomolgus (Macaca fascicularis) CD3εwith a polyhistidine affinity tag at the C-terminus (Sino Biological,Catalog 90047-C08H) was coated in 50 μl PBS (PBS, Gibco LifeTechnologies, Catalog 20012-043) onto 96-well, flat bottom plates(Maxisorb Nunc, Thermo Scientific, Catalog 12-565-226) overnight at aconcentration of 0.1 μg/ml at 4° C. The plates were washed 3 times with250 μl/well PBS-T (Tween 20 at 0.05%), blocked with 200 μl PBS-T 2% BSA(BSA Fraction V, Fisher Scientific, product BP1600-1) for 1 hour atambient temperature and washed 3 times with 250 μl/well PBS-T. Using 50μl/well, either antibody SP34scFv(LvHv)-Fc or SP34scFv(HvLv)-Fc(purified to >95% monomer) was incubated in a concentration range from37 nM to 0.017 nM diluted into PBS-T 2% BSA for 1 hour at ambienttemperature. The plates were washed three times with 250 μl/well PBS-T,incubated with the secondary anti-human Fc-HRP 1/5000 dilution intoPBS-T 2% BSA (Peroxidase AffiniPure Mouse Anti-Human IgG, Fey FragmentSpecific, Jackson ImmunoResearch, Catalog 209-035-098) for 30 minutes atambient temperature and washed 3 times with 250 μl/well PBS-T. HRPsubstrate (TMB-ELISA, Thermo Scientific, Product 34029) was added to theplate at 50 μl/well for 2 minutes and then quenched with 50 μl/well 1 MHCl before reading absorbance at 450 nM on the Tecan infinite 200Pro andanalyzed with Prism 6 (GraphPad) by curve fitting the data tolog(agonist) vs. response (three parameters). On a separate cyno CD3εcoated ELISA plate, SP34 mouse antibody (SP34-2, BD Biosciences, Catalog551916) was coated and analyzed in the same manner and using thesecondary anti-mouse IgG-HRP conjugate (Jackson ImmunoResearch, Catalog715-035-150). FIGS. 2A and 2B demonstrate that scFv antibodiesSP34scFv(LvHv)-Fc and SP34scFv(HvLv)-Fc as well as antibody SP34-2 boundto cynomolgus CD3εHIS within a similar range of EC₅₀ between 0.1 and 0.3nM.

The vectors encoding activatable antibody SP34scFv(LvHv)-Fc wereconstructed as follows. The overlapping forward primers CX2013 andCX2014 and reverse primer CX2014 were used to amplify the activatablescFv antibody using the SP34scFv(LvHv)-Fc construct as template. The DNAfragment was then cloned into the SP34scFv(LvHv)-Fc vector using theEcoR1 and BstXI restriction sites. Using the SP34scFv(LvHv)-Fc fusionvector as template, the masking peptide sequences were added using theforward primers CX2041-CX2045 for peptides 15003, 15263, 15860, 15864,and 15865, respectively, and the reverse primer CX2002 and subsequentlycloned using the SfiI and NotI restriction sites to produce nucleic acidmolecules encoding activatable anti-CD3ε antibodies.

TABLE 10  Primer sequences PRIMER NAME PRIMER SEQUENCE CX2001ACTTGTCACGAATTCGGAAGTGCAGCTGGTGGAATCTGGGGGCGGACTGGTGCAGCCT (SEQ ID NO: 392) CX2002TAGACTCGAGCGGCCGCTCATTTACCCGGAGACAGGGAGAGGCTCTT C (SEQ ID NO: 393)CX2003 TCGGCGGAGGCACCAAGCTGACCGTGCTGGGCGGCTCCCTGGACCCTAAGTCATCTGACAAAACTCACACATGCCCA (SEQ ID NO: 394) CX2004TGGGCATGTGTGAGTTTTGTCAGATGACTTAGGGTCCAGGGAGCCGCCCAGCACGGTCAGCTTGGTGCCTCCGCCGA (SEQ ID NO: 395) CX2005TGCACTTGTCACGAATTCGCAGGCTGTCGTGACACAGGAAAGCGCCCT GA (SEQ ID NO: 396)CX2006 TCTAGACTCGAGCGGCCGCTCATTTACCCGGAGACAGGGAGAGGCTCTTCT (SEQ ID NO: 397) CX2007TGGGGCCAGGGCACCCTCGTGACAGTGTCTGCTGGCGGCTCCCTGGACCCTAAGTCATCTGACAAAACTCACACATGCCCA (SEQ ID NO: 398) CX2008TGGGCATGTGTGAGTTTTGTCAGATGACTTAGGGTCCAGGGAGCCGCCAGCAGACACTGTCACGAGGGTGCCCTGGCCCCA (SEQ ID NO: 399) CX2012AGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTCAGGCTGTCGTGACACAGGAAAGCGCCCTGACCACCA (SEQ ID NO: 400) CX2013CTAAGTCTTGCACTTGTCACGAATTCGCAAGGCCAGTCTGGCCAAGGGTACCAAGGCTCGAGCGGTGGCAGCGGTGGCT (SEQ ID NO: 401) CX2014TCCGGGCCACCCATTCCAGTCCCTTGCCAGGGGCCTGGCGCACCCAATTCATGGCGTAGGTGTT (SEQ ID NO: 402) CX2041CAAGGCCAGTCTGGCCAAGGTTATCGGTGGGGTTGCGAGTGGAATTGCGGTGGGATTACTACTGGCTCGAGCGGTGGCAGCGGTGGC (SEQ ID NO: 403) CX2042CAAGGCCAGTCTGGCCAATGGTATTCGGGTGGGTGCGAGGCTTTTTGCGGTATTTTGTCGTCGGGCTCGAGCGGTGGCAGCGGTGGC (SEQ ID NO: 404) CX2043CAAGGCCAGTCTGGCCAAGTTTATTATTGCGGTGGGAATGAGAGTCTGTGCGGTGAGAGGAGGGGCTCGAGCGGTGGCAGCGGTGGC (SEQ ID NO: 405) CX2044CAAGGCCAGTCTGGCCAATTTATGTGCCAGCAGCGGATGTGGGGGAATGAGTTTTGCCATCAGGGCTCGAGCGGTGGCAGCGGTGGC (SEQ ID NO: 406) CX2045CAAGGCCAGTCTGGCCAAATGATGTATTGCGGTGGGAATGAGGTGTTGTGCGGGCCGCGGGTTGGCTCGAGCGGTGGCAGCGGTGGC (SEQ ID NO: 370)

SP34 Lv Nucleotide sequence (SEQ ID NO: 1)CAGGCTGTCGTGACACAGGAAAGCGCCCTGACCACCAGCCCTGGCGAGACAGTGACCCTGACCTGCAGATCTAGCACAGGCGCCGTGACCACAAGCAACTACGCCAACTGGGTGCAGGAAAAGCCCGACCACCTGTTCACCGGCCTGATCGGCGGCACCAACAAAAGGGCTCCAGGCGTGCCAGCCAGATTCAGCGGCAGCCTGATTGGCGATAAGGCCGCCCTGACAATCACTGGCGCCCAGACCGAGGACGAGGCCATCTACTTTTGCGCCCTGTGGTACAGCAACCTGTGGGTGTTCGGCGGAGGCACCAAGCTGACAGTGCTG  Amino Acid sequence (SEQ ID NO: 2)QAVVTQESALTTSPGETVTLTCRSSTGAVTTSNYANWVQEKPDHLFTGLIGGTNKRAPGVPARFSGSLIGDKAALTITGAQTEDEAIYFCALWYSNLWVFGGGTKLTVL  SP34 Hv Nucleotide sequence(SEQ ID NO: 3)GAAGTGCAGCTGGTGGAATCTGGCGGCGGACTGGTGCAGCCTAAGGGCTCTCTGAAGCTGAGCTGTGCCGCCAGCGGCTTCACCTTCAACACCTACGCCATGAATTGGGTGCGCCAGGCCCCTGGCAAGGGACTGGAATGGGTGGCCCGGATCAGAAGCAAGTACAACAATTACGCCACCTACTACGCCGACAGCGTGAAGGACCGGTTCACCATCAGCCGGGACGACAGCCAGAGCATCCTGTATCTGCAGATGAACAACCTGAAAACCGAGGACACCGCCATGTACTACTGCGTGCGGCACGGCAACTTCGGCAACAGCTATGTGTCTTGGTTTGCCTACTGGGGCCAGGGCACCCTCGTGACAGTGTCTGCT Amino Acid sequence (SEQ ID NO: 4)EVQLVESGGGLVQPKGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSQSILYLQMNNLKTEDTAMYYCVRHGNFGNSYVSWFAYWGQGTLVTVSA  SP34scFv(LvHv)Nucleotide sequence (SEQ ID NO: 5)CAGGCTGTCGTGACACAGGAAAGCGCCCTGACCACCAGCCCTGGCGAGACAGTGACCCTGACCTGCAGATCTAGCACAGGCGCCGTGACCACAAGCAACTACGCCAACTGGGTGCAGGAAAAGCCCGACCACCTGTTCACCGGCCTGATCGGCGGCACCAACAAAAGGGCTCCAGGCGTGCCAGCCAGATTCAGCGGCAGCCTGATTGGCGATAAGGCCGCCCTGACAATCACTGGCGCCCAGACCGAGGACGAGGCCATCTACTTTTGCGCCCTGTGGTACAGCAACCTGTGGGTGTTCGGCGGAGGCACCAAGCTGACAGTGCTGGGAGGCGGAGGATCTGGCGGAGGCGGAAGTGGCGGAGGGGGATCTGAAGTGCAGCTGGTGGAATCTGGCGGCGGACTGGTGCAGCCTAAGGGCTCTCTGAAGCTGAGCTGTGCCGCCAGCGGCTTCACCTTCAACACCTACGCCATGAATTGGGTGCGCCAGGCCCCTGGCAAGGGACTGGAATGGGTGGCCCGGATCAGAAGCAAGTACAACAATTACGCCACCTACTACGCCGACAGCGTGAAGGACCGGTTCACCATCAGCCGGGACGACAGCCAGAGCATCCTGTATCTGCAGATGAACAACCTGAAAACCGAGGACACCGCCATGTACTACTGCGTGCGGCACGGCAACTTCGGCAACAGCTATGTGTCTTGGTTTGCCTACTGGGGCCAGGGCACCCTCGTGACAGTGTCTGCT Amino Acid sequence (SEQ ID NO: 6)QAVVTQESALTTSPGETVTLTCRSSTGAVTTSNYANWVQEKPDHLFTGLIGGTNKRAPGVPARFSGSLIGDKAALTITGAQTEDEAIYFCALWYSNLWVFGGGTKLTVLGGGGSGGGGSGGGGSEVQLVESGGGLVQPKGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSQSILYLQMNNLKTEDTAMYYCVRHGNFGNSYVSWFAYWGQGTLVTVSA  Antibody SP34scFv(LvHv)-Fc fusionNucleotide sequence (SEQ ID NO: 7)CAGGCTGTCGTGACACAGGAAAGCGCCCTGACCACCAGCCCTGGCGAGACAGTGACCCTGACCTGCAGATCTAGCACAGGCGCCGTGACCACAAGCAACTACGCCAACTGGGTGCAGGAAAAGCCCGACCACCTGTTCACCGGCCTGATCGGCGGCACCAACAAAAGGGCTCCAGGCGTGCCAGCCAGATTCAGCGGCAGCCTGATTGGCGATAAGGCCGCCCTGACAATCACTGGCGCCCAGACCGAGGACGAGGCCATCTACTTTTGCGCCCTGTGGTACAGCAACCTGTGGGTGTTCGGCGGAGGCACCAAGCTGACAGTGCTGGGAGGCGGAGGATCTGGCGGAGGCGGAAGTGGCGGAGGGGGATCTGAAGTGCAGCTGGTGGAATCTGGCGGCGGACTGGTGCAGCCTAAGGGCTCTCTGAAGCTGAGCTGTGCCGCCAGCGGCTTCACCTTCAACACCTACGCCATGAATTGGGTGCGCCAGGCCCCTGGCAAGGGACTGGAATGGGTGGCCCGGATCAGAAGCAAGTACAACAATTACGCCACCTACTACGCCGACAGCGTGAAGGACCGGTTCACCATCAGCCGGGACGACAGCCAGAGCATCCTGTATCTGCAGATGAACAACCTGAAAACCGAGGACACCGCCATGTACTACTGCGTGCGGCACGGCAACTTCGGCAACAGCTATGTGTCTTGGTTTGCCTACTGGGGCCAGGGCACCCTCGTGACAGTGTCTGCTGGCGGCTCCCTGGACCCTAAGTCATCTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACAACAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAA Amino Acid sequence (SEQ ID NO: 8)QAVVTQESALTTSPGETVTLTCRSSTGAVTTSNYANWVQEKPDHLFTGLIGGTNKRAPGVPARFSGSLIGDKAALTITGAQTEDEAIYFCALWYSNLWVFGGGTKLTVLGGGGSGGGGSGGGGSEVQLVESGGGLVQPKGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSQSILYLQMNNLKTEDTAMYYCVRHGNFGNSYVSWFAYWGQGTLVTVSAGGSLDPKSSDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK  15003-1204-SP34scFv(LvHv) Nucleotide sequence[spacer (SEQ ID NO: 507)][Antibody 15003-1204-SP34scFv(LvHv)without spacer(SEQ ID NO: 547)] (SEQ ID NO: 9)[CAAGGCCAGTCTGGCCAA][GGTTATCGGTGGGGTTGCGAGTGGAATTGCGGTGGGATTACTACTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTCAGGCTGTCGTGACACAGGAAAGCGCCCTGACCACCAGCCCTGGCGAGACAGTGACCCTGACCTGCAGATCTAGCACAGGCGCCGTGACCACAAGCAACTACGCCAACTGGGTGCAGGAAAAGCCCGACCACCTGTTCACCGGCCTGATCGGCGGCACCAACAAAAGGGCTCCAGGCGTGCCAGCCAGATTCAGCGGCAGCCTGATTGGCGATAAGGCCGCCCTGACAATCACTGGCGCCCAGACCGAGGACGAGGCCATCTACTTTTGCGCCCTGTGGTACAGCAACCTGTGGGTGTTCGGCGGAGGCACCAAGCTGACAGTGCTGGGAGGCGGAGGATCTGGCGGAGGCGGAAGTGGCGGAGGGGGATCTGAAGTGCAGCTGGTGGAATCTGGCGGCGGACTGGTGCAGCCTAAGGGCTCTCTGAAGCTGAGCTGTGCCGCCAGCGGCTTCACCTTCAACACCTACGCCATGAATTGGGTGCGCCAGGCCCCTGGCAAGGGACTGGAATGGGTGGCCCGGATCAGAAGCAAGTACAACAATTACGCCACCTACTACGCCGACAGCGTGAAGGACCGGTTCACCATCAGCCGGGACGACAGCCAGAGCATCCTGTATCTGCAGATGAACAACCTGAAAACCGAGGACACCGCCATGTACTACTGCGTGCGGCACGGCAACTTCGGCAACAGCTATGTGTCTTGGTTTGCCTACTGGGGCCAGGGCACCCTCGTGACAGTGTCTGCT] Amino Acid sequence[spacer (SEQ ID NO: 87)][Antibody 15003-1204-SP34scFv(LvHv)without spacer(SEQ ID NO: 548)] (SEQ ID NO: 10)[QGQSGQ][GYRWGCEWNCGGITTGSSGGSGGSGGLSGRSDNHGGGSQAVVTQESALTTSPGETVTLTCRSSTGAVTTSNYANWVQEKPDHLFTGLIGGTNKRAPGVPARFSGSLIGDKAALTITGAQTEDEAIYFCALWYSNLWVFGGGTKLTVLGGGGSGGGGSGGGGSEVQLVESGGGLVQPKGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSQSILYLQMNNLKTEDTAMYYCVRHGNFGNSYVSWFAYWGQGTLVTVSA]Activatab le antibody 15003-1204-SP34scFv(LvHv)-Fc fusionNucleotide sequence[spacer (SEQ ID NO: 507)][Activatable Antibody 15003-1204-SP34scFv(LvHv)-Fcfusion without spacer (SEQ ID NO: 549)] (SEQ ID NO: 11)[CAAGGCCAGTCTGGCCAA][GGTTATCGGTGGGGTTGCGAGTGGAATTGCGGTGGGATTACTACTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTCAGGCTGTCGTGACACAGGAAAGCGCCCTGACCACCAGCCCTGGCGAGACAGTGACCCTGACCTGCAGATCTAGCACAGGCGCCGTGACCACAAGCAACTACGCCAACTGGGTGCAGGAAAAGCCCGACCACCTGTTCACCGGCCTGATCGGCGGCACCAACAAAAGGGCTCCAGGCGTGCCAGCCAGATTCAGCGGCAGCCTGATTGGCGATAAGGCCGCCCTGACAATCACTGGCGCCCAGACCGAGGACGAGGCCATCTACTTTTGCGCCCTGTGGTACAGCAACCTGTGGGTGTTCGGCGGAGGCACCAAGCTGACAGTGCTGGGAGGCGGAGGATCTGGCGGAGGCGGAAGTGGCGGAGGGGGATCTGAAGTGCAGCTGGTGGAATCTGGCGGCGGACTGGTGCAGCCTAAGGGCTCTCTGAAGCTGAGCTGTGCCGCCAGCGGCTTCACCTTCAACACCTACGCCATGAATTGGGTGCGCCAGGCCCCTGGCAAGGGACTGGAATGGGTGGCCCGGATCAGAAGCAAGTACAACAATTACGCCACCTACTACGCCGACAGCGTGAAGGACCGGTTCACCATCAGCCGGGACGACAGCCAGAGCATCCTGTATCTGCAGATGAACAACCTGAAAACCGAGGACACCGCCATGTACTACTGCGTGCGGCACGGCAACTTCGGCAACAGCTATGTGTCTTGGTTTGCCTACTGGGGCCAGGGCACCCTCGTGACAGTGTCTGCTGGCGGCTCCCTGGACCCTAAGTCATCTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACAACAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAA] Amino Acid Sequence[spacer (SEQ ID NO: 407)][Activatable Antibody 15003-1204-SP34scFv(LvHv)-Fcfusion without spacer (SEQ ID NO: 550)] (SEQ ID NO: 12)[QGQSGQ][GYRWGCEWNCGGITTGSSGGSGGSGGLSGRSDNHGGGSQAVVTQESALTTSPGETVTLTCRSSTGAVTTSNYANWVQEKPDHLFTGLIGGTNKRAPGVPARFSGSLIGDKAALTITGAQTEDEAIYFCALWYSNLWVFGGGTKLTVLGGGGSGGGGSGGGGSEVQLVESGGGLVQPKGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSQSILYLQMNNLKTEDTAMYYCVRHGNFGNSYVSWFAYWGQGTLVTVSAGGSLDPKSSDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]15263-1204-SP34scFv(LvHv) Nucleotide sequence[spacer (SEQ ID NO: 507)][15263-1204-SP34scFv(LvHv) without spacer(SEQ ID NO: 551)] (SEQ ID NO: 13)[CAAGGCCAGTCTGGCCAA][TGGTATTCGGGTGGGTGCGAGGCTTTTTGCGGTATTTTGTCGTCGGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTCAGGCTGTCGTGACACAGGAAAGCGCCCTGACCACCAGCCCTGGCGAGACAGTGACCCTGACCTGCAGATCTAGCACAGGCGCCGTGACCACAAGCAACTACGCCAACTGGGTGCAGGAAAAGCCCGACCACCTGTTCACCGGCCTGATCGGCGGCACCAACAAAAGGGCTCCAGGCGTGCCAGCCAGATTCAGCGGCAGCCTGATTGGCGATAAGGCCGCCCTGACAATCACTGGCGCCCAGACCGAGGACGAGGCCATCTACTTTTGCGCCCTGTGGTACAGCAACCTGTGGGTGTTCGGCGGAGGCACCAAGCTGACAGTGCTGGGAGGCGGAGGATCTGGCGGAGGCGGAAGTGGCGGAGGGGGATCTGAAGTGCAGCTGGTGGAATCTGGCGGCGGACTGGTGCAGCCTAAGGGCTCTCTGAAGCTGAGCTGTGCCGCCAGCGGCTTCACCTTCAACACCTACGCCATGAATTGGGTGCGCCAGGCCCCTGGCAAGGGACTGGAATGGGTGGCCCGGATCAGAAGCAAGTACAACAATTACGCCACCTACTACGCCGACAGCGTGAAGGACCGGTTCACCATCAGCCGGGACGACAGCCAGAGCATCCTGTATCTGCAGATGAACAACCTGAAAACCGAGGACACCGCCATGTACTACTGCGTGCGGCACGGCAACTTCGGCAACAGCTATGTGTCTTGGTTTGCCTACTGGGGCCAGGGCACCCTCGTGACAGTGTCTGCT] Amino Acid sequence[spacer (SEQ ID NO: 87)][15263-1204-SP34scFv(LvHv)without spacer(SEQ ID NO: 552)] (SEQ ID NO: 14)[QGQSGQ][WYSGGCEAFCGILSSGSSGGSGGSGGLSGRSDNHGGGSQAVVTQESALTTSPGETVTLTCRSSTGAVTTSNYANWVQEKPDHLFTGLIGGTNKRAPGVPARFSGSLIGDKAALTITGAQTEDEAIYFCALWYSNLWVFGGGTKLTVLGGGGSGGGGSGGGGSEVQLVESGGGLVQPKGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSQSILYLQMNNLKTEDTAMYYCVRHGNFGNSYVSWFAYWGQGTLVTVSA]Activatableantibody15263-1204-SP34scFv(LyHy)-FcfusionNucleotide sequence[spacer (SEQ ID NO: 507)][Activatable antibody 15263-1204-SP34scFv(LvHv)-Fcfusion without spacer (SEQ ID NO: 553)] (SEQ ID NO: 15)[CAAGGCCAGTCTGGCCAA][TGGTATTCGGGTGGGTGCGAGGCTTTTTGCGGTATTTTGTCGTCGGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTCAGGCTGTCGTGACACAGGAAAGCGCCCTGACCACCAGCCCTGGCGAGACAGTGACCCTGACCTGCAGATCTAGCACAGGCGCCGTGACCACAAGCAACTACGCCAACTGGGTGCAGGAAAAGCCCGACCACCTGTTCACCGGCCTGATCGGCGGCACCAACAAAAGGGCTCCAGGCGTGCCAGCCAGATTCAGCGGCAGCCTGATTGGCGATAAGGCCGCCCTGACAATCACTGGCGCCCAGACCGAGGACGAGGCCATCTACTTTTGCGCCCTGTGGTACAGCAACCTGTGGGTGTTCGGCGGAGGCACCAAGCTGACAGTGCTGGGAGGCGGAGGATCTGGCGGAGGCGGAAGTGGCGGAGGGGGATCTGAAGTGCAGCTGGTGGAATCTGGCGGCGGACTGGTGCAGCCTAAGGGCTCTCTGAAGCTGAGCTGTGCCGCCAGCGGCTTCACCTTCAACACCTACGCCATGAATTGGGTGCGCCAGGCCCCTGGCAAGGGACTGGAATGGGTGGCCCGGATCAGAAGCAAGTACAACAATTACGCCACCTACTACGCCGACAGCGTGAAGGACCGGTTCACCATCAGCCGGGACGACAGCCAGAGCATCCTGTATCTGCAGATGAACAACCTGAAAACCGAGGACACCGCCATGTACTACTGCGTGCGGCACGGCAACTTCGGCAACAGCTATGTGTCTTGGTTTGCCTACTGGGGCCAGGGCACCCTCGTGACAGTGTCTGCTGGCGGCTCCCTGGACCCTAAGTCATCTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACAACAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAA] Amino acid sequence[spacer (SEQ ID NO: 87)][Activatable antibody 15263-1204-SP34scFv(LvHv)-Fcfusion without spacer (SEQ ID NO: 554)]] (SEQ ID NO: 16)[QGQSGQ][WYSGGCEAFCGILSSGSSGGSGGSGGLSGRSDNHGGGSQAVVTQESALTTSPGETVTLTCRSSTGAVTTSNYANWVQEKPDHLFTGLIGGTNKRAPGVPARFSGSLIGDKAALTITGAQTEDEAIYFCALWYSNLWVFGGGTKLTVLGGGGSGGGGSGGGGSEVQLVESGGGLVQPKGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSQSILYLQMNNLKTEDTAMYYCVRHGNFGNSYVSWFAYWGQGTLVTVSAGGSLDPKSSDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]15860-1204-SP34scFv(LvHv) Nucleotide sequence[spacer (SEQ ID NO: 507)][15860-1204-SP34scFv(LvHv) without spacer(SEQ ID NO: 555)] (SEQ ID NO: 17)[CAAGGCCAGTCTGGCCAA][GTTTATTATTGCGGTGGGAATGAGAGTCTGTGCGGTGAGAGGAGGGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTCAGGCTGTCGTGACACAGGAAAGCGCCCTGACCACCAGCCCTGGCGAGACAGTGACCCTGACCTGCAGATCTAGCACAGGCGCCGTGACCACAAGCAACTACGCCAACTGGGTGCAGGAAAAGCCCGACCACCTGTTCACCGGCCTGATCGGCGGCACCAACAAAAGGGCTCCAGGCGTGCCAGCCAGATTCAGCGGCAGCCTGATTGGCGATAAGGCCGCCCTGACAATCACTGGCGCCCAGACCGAGGACGAGGCCATCTACTTTTGCGCCCTGTGGTACAGCAACCTGTGGGTGTTCGGCGGAGGCACCAAGCTGACAGTGCTGGGAGGCGGAGGATCTGGCGGAGGCGGAAGTGGCGGAGGGGGATCTGAAGTGCAGCTGGTGGAATCTGGCGGCGGACTGGTGCAGCCTAAGGGCTCTCTGAAGCTGAGCTGTGCCGCCAGCGGCTTCACCTTCAACACCTACGCCATGAATTGGGTGCGCCAGGCCCCTGGCAAGGGACTGGAATGGGTGGCCCGGATCAGAAGCAAGTACAACAATTACGCCACCTACTACGCCGACAGCGTGAAGGACCGGTTCACCATCAGCCGGGACGACAGCCAGAGCATCCTGTATCTGCAGATGAACAACCTGAAAACCGAGGACACCGCCATGTACTACTGCGTGCGGCACGGCAACTTCGGCAACAGCTATGTGTCTTGGTTTGCCTACTGGGGCCAGGGCACCCTCGTGACAGTGTCTGCT] Amino Acid sequence[spacer (SEQ ID NO: 87)][15263-1204-SP34scFv(LvHv)without spacer(SEQ ID NO: 556)] (SEQ ID NO: 18)[QGQSGQ][VYYCGGNESLCGERRGSSGGSGGSGGLSGRSDNHGGGSQAVVTQESALTTSPGETVTLTCRSSTGAVTTSNYANWVQEKPDHLFTGLIGGTNKRAPGVPARFSGSLIGDKAALTITGAQTEDEAIYFCALWYSNLWVFGGGTKLTVLGGGGSGGGGSGGGGSEVQLVESGGGLVQPKGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSQSILYLQMNNLKTEDTAMYYCVRHGNFGNSYVSWFAYWGQGTLVTVSA]Activatable antibody 15860-1204-SP34scFv(LvHv)-Fc fusionNucleotide sequence[spacer (SEQ ID NO: 507)][Activatable antibody 15860-1204-SP34scFv(LvHv)-Fcfusion without spacer (SEQ ID NO: 557)] (SEQ ID NO: 19)[CAAGGCCAGTCTGGCCAA][GTTTATTATTGCGGTGGGAATGAGAGTCTGTGCGGTGAGAGGAGGGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTCAGGCTGTCGTGACACAGGAAAGCGCCCTGACCACCAGCCCTGGCGAGACAGTGACCCTGACCTGCAGATCTAGCACAGGCGCCGTGACCACAAGCAACTACGCCAACTGGGTGCAGGAAAAGCCCGACCACCTGTTCACCGGCCTGATCGGCGGCACCAACAAAAGGGCTCCAGGCGTGCCAGCCAGATTCAGCGGCAGCCTGATTGGCGATAAGGCCGCCCTGACAATCACTGGCGCCCAGACCGAGGACGAGGCCATCTACTTTTGCGCCCTGTGGTACAGCAACCTGTGGGTGTTCGGCGGAGGCACCAAGCTGACAGTGCTGGGAGGCGGAGGATCTGGCGGAGGCGGAAGTGGCGGAGGGGGATCTGAAGTGCAGCTGGTGGAATCTGGCGGCGGACTGGTGCAGCCTAAGGGCTCTCTGAAGCTGAGCTGTGCCGCCAGCGGCTTCACCTTCAACACCTACGCCATGAATTGGGTGCGCCAGGCCCCTGGCAAGGGACTGGAATGGGTGGCCCGGATCAGAAGCAAGTACAACAATTACGCCACCTACTACGCCGACAGCGTGAAGGACCGGTTCACCATCAGCCGGGACGACAGCCAGAGCATCCTGTATCTGCAGATGAACAACCTGAAAACCGAGGACACCGCCATGTACTACTGCGTGCGGCACGGCAACTTCGGCAACAGCTATGTGTCTTGGTTTGCCTACTGGGGCCAGGGCACCCTCGTGACAGTGTCTGCTGGCGGCTCCCTGGACCCTAAGTCATCTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACAACAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAA] Amino Acid sequence[spacer (SEQ ID NO: 87)][Activatable antibody 15860-1204-SP34scFv(LvHv)-Fcfusion without spacer (SEQ ID NO: 558)] (SEQ ID NO: 20)[QGQSGQ][VYYCGGNESLCGERRGSSGGSGGSGGLSGRSDNHGGGSQAVVTQESALTTSPGETVTLTCRSSTGAVTTSNYANWVQEKPDHLFTGLIGGTNKRAPGVPARFSGSLIGDKAALTITGAQTEDEAIYFCALWYSNLWVFGGGTKLTVLGGGGSGGGGSGGGGSEVQLVESGGGLVQPKGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSQSILYLQMNNLKTEDTAMYYCVRHGNFGNSYVSWFAYWGQGTLVTVSAGGSLDPKSSDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]15864-1204-SP34scFv(LvHv) Nucleotide sequence[spacer (SEQ ID NO: 507)][15864-1204-SP34scFv(LvHv) without spacer (SEQ IDNO: 559)] (SEQ ID NO: 21)[CAAGGCCAGTCTGGCCAA][TTTATGTGCCAGCAGCGGATGTGGGGGAATGAGTTTTGCCATCAGGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTCAGGCTGTCGTGACACAGGAAAGCGCCCTGACCACCAGCCCTGGCGAGACAGTGACCCTGACCTGCAGATCTAGCACAGGCGCCGTGACCACAAGCAACTACGCCAACTGGGTGCAGGAAAAGCCCGACCACCTGTTCACCGGCCTGATCGGCGGCACCAACAAAAGGGCTCCAGGCGTGCCAGCCAGATTCAGCGGCAGCCTGATTGGCGATAAGGCCGCCCTGACAATCACTGGCGCCCAGACCGAGGACGAGGCCATCTACTTTTGCGCCCTGTGGTACAGCAACCTGTGGGTGTTCGGCGGAGGCACCAAGCTGACAGTGCTGGGAGGCGGAGGATCTGGCGGAGGCGGAAGTGGCGGAGGGGGATCTGAAGTGCAGCTGGTGGAATCTGGCGGCGGACTGGTGCAGCCTAAGGGCTCTCTGAAGCTGAGCTGTGCCGCCAGCGGCTTCACCTTCAACACCTACGCCATGAATTGGGTGCGCCAGGCCCCTGGCAAGGGACTGGAATGGGTGGCCCGGATCAGAAGCAAGTACAACAATTACGCCACCTACTACGCCGACAGCGTGAAGGACCGGTTCACCATCAGCCGGGACGACAGCCAGAGCATCCTGTATCTGCAGATGAACAACCTGAAAACCGAGGACACCGCCATGTACTACTGCGTGCGGCACGGCAACTTCGGCAACAGCTATGTGTCTTGGTTTGCCTACTGGGGCCAGGGCACCCTCGTGACAGTGTCTGCT] Amino Acid[spacer (SEQ ID NO: 87)][15864-1204-SP34scFv(LvHv) without spacer (SEQ IDNO: 560)] (SEQ ID NO: 22)[QGQSGQ][FMCQQRMWGNEFCHQGSSGGSGGSGGLSGRSDNHGGGSQAVVTQESALTTSPGETVTLTCRSSTGAVTTSNYANWVQEKPDHLFTGLIGGTNKRAPGVPARFSGSLIGDKAALTITGAQTEDEAIYFCALWYSNLWVFGGGTKLTVLGGGGSGGGGSGGGGSEVQLVESGGGLVQPKGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSQSILYLQMNNLKTEDTAMYYCVRHGNFGNSYVSWFAYWGQGTLVTVSA]Activatab le antibody 15864-1204-SP34scFv(Lvtiv)-Fc fusionNucleotide sequence[spacer (SEQ ID NO: 507)][Activatable antibody 15864-1204-SP34scFv(LvHv)-Fcfusion without spacer (SEQ ID NO: 561)] (SEQ ID NO: 23)[CAAGGCCAGTCTGGCCAA][TTTATGTGCCAGCAGCGGATGTGGGGGAATGAGTTTTGCCATCAGGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTCAGGCTGTCGTGACACAGGAAAGCGCCCTGACCACCAGCCCTGGCGAGACAGTGACCCTGACCTGCAGATCTAGCACAGGCGCCGTGACCACAAGCAACTACGCCAACTGGGTGCAGGAAAAGCCCGACCACCTGTTCACCGGCCTGATCGGCGGCACCAACAAAAGGGCTCCAGGCGTGCCAGCCAGATTCAGCGGCAGCCTGATTGGCGATAAGGCCGCCCTGACAATCACTGGCGCCCAGACCGAGGACGAGGCCATCTACTTTTGCGCCCTGTGGTACAGCAACCTGTGGGTGTTCGGCGGAGGCACCAAGCTGACAGTGCTGGGAGGCGGAGGATCTGGCGGAGGCGGAAGTGGCGGAGGGGGATCTGAAGTGCAGCTGGTGGAATCTGGCGGCGGACTGGTGCAGCCTAAGGGCTCTCTGAAGCTGAGCTGTGCCGCCAGCGGCTTCACCTTCAACACCTACGCCATGAATTGGGTGCGCCAGGCCCCTGGCAAGGGACTGGAATGGGTGGCCCGGATCAGAAGCAAGTACAACAATTACGCCACCTACTACGCCGACAGCGTGAAGGACCGGTTCACCATCAGCCGGGACGACAGCCAGAGCATCCTGTATCTGCAGATGAACAACCTGAAAACCGAGGACACCGCCATGTACTACTGCGTGCGGCACGGCAACTTCGGCAACAGCTATGTGTCTTGGTTTGCCTACTGGGGCCAGGGCACCCTCGTGACAGTGTCTGCTGGCGGCTCCCTGGACCCTAAGTCATCTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACAACAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAA] Amino acid sequence[spacer (SEQ ID NO: 87)][Activatable antibody 15864-1204-SP34scFv(LvHv)-Fcfusion without spacer (SEQ ID NO: 562)] (SEQ ID NO: 24)[QGQSGQ][FMCQQRMWGNEFCHQGSSGGSGGSGGLSGRSDNHGGGSQAVVTQESALTTSPGETVTLTCRSSTGAVTTSNYANWVQEKPDHLFTGLIGGTNKRAPGVPARFSGSLIGDKAALTITGAQTEDEAIYFCALWYSNLWVFGGGTKLTVLGGGGSGGGGSGGGGSEVQLVESGGGLVQPKGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSQSILYLQMNNLKTEDTAMYYCVRHGNFGNSYVSWFAYWGQGTLVTVSAGGSLDPKSSDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]15865-1204-SP34scFv(LvHv) Nucleotide sequence[spacer (SEQ ID NO: 507)][15865-1204-SP34scFv(LvHv) without spacer (SEQ IDNO: 563)] (SEQ ID NO: 25)[CAAGGCCAGTCTGGCCAA][ATGATGTATTGCGGTGGGAATGAGGTGTTGTGCGGGCCGCGGGTTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTCAGGCTGTCGTGACACAGGAAAGCGCCCTGACCACCAGCCCTGGCGAGACAGTGACCCTGACCTGCAGATCTAGCACAGGCGCCGTGACCACAAGCAACTACGCCAACTGGGTGCAGGAAAAGCCCGACCACCTGTTCACCGGCCTGATCGGCGGCACCAACAAAAGGGCTCCAGGCGTGCCAGCCAGATTCAGCGGCAGCCTGATTGGCGATAAGGCCGCCCTGACAATCACTGGCGCCCAGACCGAGGACGAGGCCATCTACTTTTGCGCCCTGTGGTACAGCAACCTGTGGGTGTTCGGCGGAGGCACCAAGCTGACAGTGCTGGGAGGCGGAGGATCTGGCGGAGGCGGAAGTGGCGGAGGGGGATCTGAAGTGCAGCTGGTGGAATCTGGCGGCGGACTGGTGCAGCCTAAGGGCTCTCTGAAGCTGAGCTGTGCCGCCAGCGGCTTCACCTTCAACACCTACGCCATGAATTGGGTGCGCCAGGCCCCTGGCAAGGGACTGGAATGGGTGGCCCGGATCAGAAGCAAGTACAACAATTACGCCACCTACTACGCCGACAGCGTGAAGGACCGGTTCACCATCAGCCGGGACGACAGCCAGAGCATCCTGTATCTGCAGATGAACAACCTGAAAACCGAGGACACCGCCATGTACTACTGCGTGCGGCACGGCAACTTCGGCAACAGCTATGTGTCTTGGTTTGCCTACTGGGGCCAGGGCACCCTCGTGACAGTGTCTGCT] Amino Acid sequence[spacer (SEQ ID NO: 87)][15865-1204-SP34scFv(LvHv) without spacer (SEQ IDNO: 564)] (SEQ ID NO: 26)[QGQSGQ][MMYCGGNEVLCGPRVGSSGGSGGSGGLSGRSDNHGGGSQAVVTQESALTTSPGETVTLTCRSSTGAVTTSNYANWVQEKPDHLFTGLIGGTNKRAPGVPARFSGSLIGDKAALTITGAQTEDEAIYFCALWYSNLWVFGGGTKLTVLGGGGSGGGGSGGGGSEVQLVESGGGLVQPKGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSQSILYLQMNNLKTEDTAMYYCVRHGNFGNSYVSWFAYWGQGTLVTVSA]Activatable antibody 15865-1204-SP34scFv(LvHv)-Fc fusionNucleotide sequence[spacer (SEQ ID NO: 507)][Activatable antibody 15865-1204-SP34scFv(LvHv)-Fcfusion without spacer (SEQ ID NO: 565)] (SEQ ID NO: 27)[CAAGGCCAGTCTGGCCAA][ATGATGTATTGCGGTGGGAATGAGGTGTTGTGCGGGCCGCGGGTTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTCAGGCTGTCGTGACACAGGAAAGCGCCCTGACCACCAGCCCTGGCGAGACAGTGACCCTGACCTGCAGATCTAGCACAGGCGCCGTGACCACAAGCAACTACGCCAACTGGGTGCAGGAAAAGCCCGACCACCTGTTCACCGGCCTGATCGGCGGCACCAACAAAAGGGCTCCAGGCGTGCCAGCCAGATTCAGCGGCAGCCTGATTGGCGATAAGGCCGCCCTGACAATCACTGGCGCCCAGACCGAGGACGAGGCCATCTACTTTTGCGCCCTGTGGTACAGCAACCTGTGGGTGTTCGGCGGAGGCACCAAGCTGACAGTGCTGGGAGGCGGAGGATCTGGCGGAGGCGGAAGTGGCGGAGGGGGATCTGAAGTGCAGCTGGTGGAATCTGGCGGCGGACTGGTGCAGCCTAAGGGCTCTCTGAAGCTGAGCTGTGCCGCCAGCGGCTTCACCTTCAACACCTACGCCATGAATTGGGTGCGCCAGGCCCCTGGCAAGGGACTGGAATGGGTGGCCCGGATCAGAAGCAAGTACAACAATTACGCCACCTACTACGCCGACAGCGTGAAGGACCGGTTCACCATCAGCCGGGACGACAGCCAGAGCATCCTGTATCTGCAGATGAACAACCTGAAAACCGAGGACACCGCCATGTACTACTGCGTGCGGCACGGCAACTTCGGCAACAGCTATGTGTCTTGGTTTGCCTACTGGGGCCAGGGCACCCTCGTGACAGTGTCTGCTGGCGGCTCCCTGGACCCTAAGTCATCTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACAACAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAA] Amino Acid sequence[spacer (SEQ ID NO: 87)][Activatable antibody 15865-1204-SP34scFv(LvHv)-Fcfusion without spacer (SEQ ID NO: 566)] (SEQ ID NO: 28)[QGQSGQ][MMYCGGNEVLCGPRVGSSGGSGGSGGLSGRSDNHGGGSQAVVTQESALTTSPGETVTLTCRSSTGAVTTSNYANWVQEKPDHLFTGLIGGTNKRAPGVPARFSGSLIGDKAALTITGAQTEDEAIYFCALWYSNLWVFGGGTKLTVLGGGGSGGGGSGGGGSEVQLVESGGGLVQPKGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSQSILYLQMNNLKTEDTAMYYCVRHGNFGNSYVSWFAYWGQGTLVTVSAGGSLDPKSSDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]SP34scFv(HvLv) Nucleotide sequence (SEQ ID NO: 29)GAAGTGCAGCTGGTGGAATCTGGGGGCGGACTGGTGCAGCCTAAGGGCAGCCTGAAGCTGAGCTGTGCCGCCAGCGGCTTCACCTTCAACACCTACGCCATGAACTGGGTGCGCCAGGCCCCTGGCAAAGGCCTGGAATGGGTGGCCCGGATCAGAAGCAAGTACAACAATTACGCCACCTACTACGCCGACAGCGTGAAGGACAGATTCACCATCAGCCGGGACGACAGCCAGAGCATCCTGTACCTGCAGATGAACAACCTGAAAACCGAGGACACCGCCATGTACTACTGCGTGCGGCACGGCAACTTCGGCAACAGCTATGTGTCTTGGTTTGCCTACTGGGGCCAGGGCACCCTCGTGACAGTGTCTGCCGGTGGTGGTGGTAGTGGTGGCGGTGGTTCAGGCGGTGGCGGTAGCCAGGCTGTCGTGACACAGGAAAGCGCCCTGACCACCAGCCCTGGCGAGACAGTGACCCTGACCTGTAGAAGCAGCACAGGCGCCGTGACCACAAGCAACTACGCCAATTGGGTGCAGGAAAAGCCCGACCACCTGTTCACCGGCCTGATCGGCGGCACCAACAAAAGGGCTCCAGGCGTGCCAGCCAGATTCAGCGGCTCCCTGATCGGAGATAAGGCCGCCCTGACAATCACTGGCGCCCAGACCGAGGACGAGGCCATCTACTTTTGCGCCCTGTGGTACAGCAACCTGTGGGTGTTCGGCGGAGGCACCAAGCTGACCGTGCTG Amino Acid sequence (SEQ ID NO: 30)EVQLVESGGGLVQPKGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSQSILYLQMNNLKTEDTAMYYCVRHGNFGNSYVSWFAYWGQGTLVTVSAGGGGSGGGGSGGGGSQAVVTQESALTTSPGETVTLTCRSSTGAVTTSNYANWVQEKPDHLFTGLIGGTNKRAPGVPARFSGSLIGDKAALTITGAQTEDEAIYFCALWYSNLWVFGGGTKLTVL  Antibody SP34scFv(HvLv)-Fc fusionNucleotide sequence (SEQ ID NO: 31)GAAGTGCAGCTGGTGGAATCTGGGGGCGGACTGGTGCAGCCTAAGGGCAGCCTGAAGCTGAGCTGTGCCGCCAGCGGCTTCACCTTCAACACCTACGCCATGAACTGGGTGCGCCAGGCCCCTGGCAAAGGCCTGGAATGGGTGGCCCGGATCAGAAGCAAGTACAACAATTACGCCACCTACTACGCCGACAGCGTGAAGGACAGATTCACCATCAGCCGGGACGACAGCCAGAGCATCCTGTACCTGCAGATGAACAACCTGAAAACCGAGGACACCGCCATGTACTACTGCGTGCGGCACGGCAACTTCGGCAACAGCTATGTGTCTTGGTTTGCCTACTGGGGCCAGGGCACCCTCGTGACAGTGTCTGCCGGTGGTGGTGGTAGTGGTGGCGGTGGTTCAGGCGGTGGCGGTAGCCAGGCTGTCGTGACACAGGAAAGCGCCCTGACCACCAGCCCTGGCGAGACAGTGACCCTGACCTGTAGAAGCAGCACAGGCGCCGTGACCACAAGCAACTACGCCAATTGGGTGCAGGAAAAGCCCGACCACCTGTTCACCGGCCTGATCGGCGGCACCAACAAAAGGGCTCCAGGCGTGCCAGCCAGATTCAGCGGCTCCCTGATCGGAGATAAGGCCGCCCTGACAATCACTGGCGCCCAGACCGAGGACGAGGCCATCTACTTTTGCGCCCTGTGGTACAGCAACCTGTGGGTGTTCGGCGGAGGCACCAAGCTGACCGTGCTGGGCGGCTCCCTGGACCCTAAGTCATCTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACAACAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAA Amino Acid sequence (SEQ ID NO: 32)EVQLVESGGGLVQPKGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSQSILYLQMNNLKTEDTAMYYCVRHGNFGNSYVSWFAYWGQGTLVTVSAGGGGSGGGGSGGGGSQAVVTQESALTTSPGETVTLTCRSSTGAVTTSNYANWVQEKPDHLFTGLIGGTNKRAPGVPARFSGSLIGDKAALTITGAQTEDEAIYFCALWYSNLWVFGGGTKLTVLGGSLDPKSSDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK 15003-1204-SP34scFv(HvLv) Nucleotide sequence[spacer (SEQ ID NO: 507)][15003-1204-SP34scFv(HvLv) without spacer (SEQ IDNO: 567)] (SEQ ID NO: 33)[CAAGGCCAGTCTGGCCAA][GGTTATCGGTGGGGTTGCGAGTGGAATTGCGGTGGGATTACTACTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTGAAGTGCAGCTGGTGGAATCTGGGGGCGGACTGGTGCAGCCTAAGGGCAGCCTGAAGCTGAGCTGTGCCGCCAGCGGCTTCACCTTCAACACCTACGCCATGAACTGGGTGCGCCAGGCCCCTGGCAAAGGCCTGGAATGGGTGGCCCGGATCAGAAGCAAGTACAACAATTACGCCACCTACTACGCCGACAGCGTGAAGGACAGATTCACCATCAGCCGGGACGACAGCCAGAGCATCCTGTACCTGCAGATGAACAACCTGAAAACCGAGGACACCGCCATGTACTACTGCGTGCGGCACGGCAACTTCGGCAACAGCTATGTGTCTTGGTTTGCCTACTGGGGCCAGGGCACCCTCGTGACAGTGTCTGCCGGTGGTGGTGGTAGTGGTGGCGGTGGTTCAGGCGGTGGCGGTAGCCAGGCTGTCGTGACACAGGAAAGCGCCCTGACCACCAGCCCTGGCGAGACAGTGACCCTGACCTGTAGAAGCAGCACAGGCGCCGTGACCACAAGCAACTACGCCAATTGGGTGCAGGAAAAGCCCGACCACCTGTTCACCGGCCTGATCGGCGGCACCAACAAAAGGGCTCCAGGCGTGCCAGCCAGATTCAGCGGCTCCCTGATCGGAGATAAGGCCGCCCTGACAATCACTGGCGCCCAGACCGAGGACGAGGCCATCTACTTTTGCGCCCTGTGGTACAGCAACCTGTGGGTGTTCGGCGGAGGCACCAAGCTGACCGTGCTG  Amino Acid sequence[spacer (SEQ ID NO: 87)][15003-1204-SP34scFv(HvLv)without spacer (SEQ IDNO: 568)] (SEQ ID NO: 34)[QGQSGQ][GYRWGCEWNCGGITTGSSGGSGGSGGLSGRSDNHGGGSEVQLVESGGGLVQPKGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSQSILYLQMNNLKTEDTAMYYCVRHGNFGNSYVSWFAYWGQGTLVTVSAGGGGSGGGGSGGGGSQAVVTQESALTTSPGETVTLTCRSSTGAVTTSNYANWVQEKPDHLFTGLIGGTNKRAPGVPARFSGSLIGDKAALTITGAQTEDEAIYFCALWYSNLWVFGGGTKLTVL]Activatable antibody 15003-1204-SP34scFv(HvLv)-Fc fusionNucleotide sequence[spacer (SEQ ID NO: 507)][Activatable antibody 15003-1204-SP34scFv(HvLv)-Fcfusion without spacer (SEQ ID NO: 569)] (SEQ ID NO: 35)[CAAGGCCAGTCTGGCCAA][GGTTATCGGTGGGGTTGCGAGTGGAATTGCGGTGGGATTACTACTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTGAAGTGCAGCTGGTGGAATCTGGGGGCGGACTGGTGCAGCCTAAGGGCAGCCTGAAGCTGAGCTGTGCCGCCAGCGGCTTCACCTTCAACACCTACGCCATGAACTGGGTGCGCCAGGCCCCTGGCAAAGGCCTGGAATGGGTGGCCCGGATCAGAAGCAAGTACAACAATTACGCCACCTACTACGCCGACAGCGTGAAGGACAGATTCACCATCAGCCGGGACGACAGCCAGAGCATCCTGTACCTGCAGATGAACAACCTGAAAACCGAGGACACCGCCATGTACTACTGCGTGCGGCACGGCAACTTCGGCAACAGCTATGTGTCTTGGTTTGCCTACTGGGGCCAGGGCACCCTCGTGACAGTGTCTGCCGGTGGTGGTGGTAGTGGTGGCGGTGGTTCAGGCGGTGGCGGTAGCCAGGCTGTCGTGACACAGGAAAGCGCCCTGACCACCAGCCCTGGCGAGACAGTGACCCTGACCTGTAGAAGCAGCACAGGCGCCGTGACCACAAGCAACTACGCCAATTGGGTGCAGGAAAAGCCCGACCACCTGTTCACCGGCCTGATCGGCGGCACCAACAAAAGGGCTCCAGGCGTGCCAGCCAGATTCAGCGGCTCCCTGATCGGAGATAAGGCCGCCCTGACAATCACTGGCGCCCAGACCGAGGACGAGGCCATCTACTTTTGCGCCCTGTGGTACAGCAACCTGTGGGTGTTCGGCGGAGGCACCAAGCTGACCGTGCTGGGCGGCTCCCTGGACCCTAAGTCATCTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACAACAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAA  Amino Acid sequence[spacer (SEQ ID NO: 87)][Activatable antibody 15003-1204-SP34scFv(HvLv)-Fcfusion without spacer (SEQ ID NO: 570)] (SEQ ID NO: 36)[QGQSGQ][GYRWGCEWNCGGITTGSSGGSGGSGGLSGRSDNHGGGSEVQLVESGGGLVQPKGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSQSILYLQMNNLKTEDTAMYYCVRHGNFGNSYVSWFAYWGQGTLVTVSAGGGGSGGGGSGGGGSQAVVTQESALTTSPGETVTLTCRSSTGAVTTSNYANWVQEKPDHLFTGLIGGTNKRAPGVPARFSGSLIGDKAALTITGAQTEDEAIYFCALWYSNLWVFGGGTKLTVLGGSLDPKSSDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]15263-1204-SP34scFv(HvLv) Nucleotide sequence[spacer (SEQ ID NO: 507)][15263-1204-SP34scFv(HvLv) without spacer (SEQ IDNO: 571)] (SEQ ID NO: 37)[CAAGGCCAGTCTGGCCAA][TGGTATTCGGGTGGGTGCGAGGCTTTTTGCGGTATTTTGTCGTCGGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTGAAGTGCAGCTGGTGGAATCTGGGGGCGGACTGGTGCAGCCTAAGGGCAGCCTGAAGCTGAGCTGTGCCGCCAGCGGCTTCACCTTCAACACCTACGCCATGAACTGGGTGCGCCAGGCCCCTGGCAAAGGCCTGGAATGGGTGGCCCGGATCAGAAGCAAGTACAACAATTACGCCACCTACTACGCCGACAGCGTGAAGGACAGATTCACCATCAGCCGGGACGACAGCCAGAGCATCCTGTACCTGCAGATGAACAACCTGAAAACCGAGGACACCGCCATGTACTACTGCGTGCGGCACGGCAACTTCGGCAACAGCTATGTGTCTTGGTTTGCCTACTGGGGCCAGGGCACCCTCGTGACAGTGTCTGCCGGTGGTGGTGGTAGTGGTGGCGGTGGTTCAGGCGGTGGCGGTAGCCAGGCTGTCGTGACACAGGAAAGCGCCCTGACCACCAGCCCTGGCGAGACAGTGACCCTGACCTGTAGAAGCAGCACAGGCGCCGTGACCACAAGCAACTACGCCAATTGGGTGCAGGAAAAGCCCGACCACCTGTTCACCGGCCTGATCGGCGGCACCAACAAAAGGGCTCCAGGCGTGCCAGCCAGATTCAGCGGCTCCCTGATCGGAGATAAGGCCGCCCTGACAATCACTGGCGCCCAGACCGAGGACGAGGCCATCTACTTTTGCGCCCTGTGGTACAGCAACCTGTGGGTGTTCGGCGGAGGCACCAAGCTGACCGTGCTG] Amino Acid sequence[spacer (SEQ ID NO: 87)][15263-1204-SP34scFv(HvLv) without spacer (SEQ IDNO: 572)] (SEQ ID NO: 38)[QGQSGQ][WYSGGCEAFCGILSSGSSGGSGGSGGLSGRSDNHGGGSEVQLVESGGGLVQPKGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSQSILYLQMNNLKTEDTAMYYCVRHGNFGNSYVSWFAYWGQGTLVTVSAGGGGSGGGGSGGGGSQAVVTQESALTTSPGETVTLTCRSSTGAVTTSNYANWVQEKPDHLFTGLIGGTNKRAPGVPARFSGSLIGDKAALTITGAQTEDEAIYFCALWYSNLWVFGGGTKLTVL]Activatable antibody 15263-1204-SP34scFv(HvLv)-Fc fusionNucleotide sequence[spacer (SEQ ID NO: 507)][Activatable antibody 15263-1204-SP34scFv(HvLv)-Fcfusion without spacer (SEQ ID NO: 573)] (SEQ ID NO: 39)[CAAGGCCAGTCTGGCCAA][TGGTATTCGGGTGGGTGCGAGGCTTTTTGCGGTATTTTGTCGTCGGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTGAAGTGCAGCTGGTGGAATCTGGGGGCGGACTGGTGCAGCCTAAGGGCAGCCTGAAGCTGAGCTGTGCCGCCAGCGGCTTCACCTTCAACACCTACGCCATGAACTGGGTGCGCCAGGCCCCTGGCAAAGGCCTGGAATGGGTGGCCCGGATCAGAAGCAAGTACAACAATTACGCCACCTACTACGCCGACAGCGTGAAGGACAGATTCACCATCAGCCGGGACGACAGCCAGAGCATCCTGTACCTGCAGATGAACAACCTGAAAACCGAGGACACCGCCATGTACTACTGCGTGCGGCACGGCAACTTCGGCAACAGCTATGTGTCTTGGTTTGCCTACTGGGGCCAGGGCACCCTCGTGACAGTGTCTGCCGGTGGTGGTGGTAGTGGTGGCGGTGGTTCAGGCGGTGGCGGTAGCCAGGCTGTCGTGACACAGGAAAGCGCCCTGACCACCAGCCCTGGCGAGACAGTGACCCTGACCTGTAGAAGCAGCACAGGCGCCGTGACCACAAGCAACTACGCCAATTGGGTGCAGGAAAAGCCCGACCACCTGTTCACCGGCCTGATCGGCGGCACCAACAAAAGGGCTCCAGGCGTGCCAGCCAGATTCAGCGGCTCCCTGATCGGAGATAAGGCCGCCCTGACAATCACTGGCGCCCAGACCGAGGACGAGGCCATCTACTTTTGCGCCCTGTGGTACAGCAACCTGTGGGTGTTCGGCGGAGGCACCAAGCTGACCGTGCTGGGCGGCTCCCTGGACCCTAAGTCATCTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACAACAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAA] Amino Acid sequence[spacer (SEQ ID NO: 87)][Activatable antibody 15263-1204-SP34scFv(HvLv)-Fcfusion without spacer (SEQ ID NO: 574)] (SEQ ID NO: 40)[QGQSGQ][WYSGGCEAFCGILSSGSSGGSGGSGGLSGRSDNHGGGSEVQLVESGGGLVQPKGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSQSILYLQMNNLKTEDTAMYYCVRHGNFGNSYVSWFAYWGQGTLVTVSAGGGGSGGGGSGGGGSQAVVTQESALTTSPGETVTLTCRSSTGAVTTSNYANWVQEKPDHLFTGLIGGTNKRAPGVPARFSGSLIGDKAALTITGAQTEDEAIYFCALWYSNLWVFGGGTKLTVLGGSLDPKSSDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]15860-1204-SP34scFv(HvLv) Nucleotide sequence[spacer (SEQ ID NO: 507)][15860-1204-SP34scFv(HvLv) without spacer (SEQ IDNO: 575)] (SEQ ID NO: 41)[CAAGGCCAGTCTGGCCAA][GTTTATTATTGCGGTGGGAATGAGAGTCTGTGCGGTGAGAGGAGGGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTGAAGTGCAGCTGGTGGAATCTGGGGGCGGACTGGTGCAGCCTAAGGGCAGCCTGAAGCTGAGCTGTGCCGCCAGCGGCTTCACCTTCAACACCTACGCCATGAACTGGGTGCGCCAGGCCCCTGGCAAAGGCCTGGAATGGGTGGCCCGGATCAGAAGCAAGTACAACAATTACGCCACCTACTACGCCGACAGCGTGAAGGACAGATTCACCATCAGCCGGGACGACAGCCAGAGCATCCTGTACCTGCAGATGAACAACCTGAAAACCGAGGACACCGCCATGTACTACTGCGTGCGGCACGGCAACTTCGGCAACAGCTATGTGTCTTGGTTTGCCTACTGGGGCCAGGGCACCCTCGTGACAGTGTCTGCCGGTGGTGGTGGTAGTGGTGGCGGTGGTTCAGGCGGTGGCGGTAGCCAGGCTGTCGTGACACAGGAAAGCGCCCTGACCACCAGCCCTGGCGAGACAGTGACCCTGACCTGTAGAAGCAGCACAGGCGCCGTGACCACAAGCAACTACGCCAATTGGGTGCAGGAAAAGCCCGACCACCTGTTCACCGGCCTGATCGGCGGCACCAACAAAAGGGCTCCAGGCGTGCCAGCCAGATTCAGCGGCTCCCTGATCGGAGATAAGGCCGCCCTGACAATCACTGGCGCCCAGACCGAGGACGAGGCCATCTACTTTTGCGCCCTGTGGTACAGCAACCTGTGGGTGTTCGGCGGAGGCACCAAGCTGACCGTGCTG] Amino Acid sequence[spacer (SEQ ID NO: 87)][15860-1204-SP34scFv(HvLv) without spacer (SEQ IDNO: 576)] (SEQ ID NO: 42)[QGQSGQ][VYYCGGNESLCGERRGSSGGSGGSGGLSGRSDNHGGGSEVQLVESGGGLVQPKGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSQSILYLQMNNLKTEDTAMYYCVRHGNFGNSYVSWFAYWGQGTLVTVSAGGGGSGGGGSGGGGSQAVVTQESALTTSPGETVTLTCRSSTGAVTTSNYANWVQEKPDHLFTGLIGGTNKRAPGVPARFSGSLIGDKAALTITGAQTEDEAIYFCALWYSNLWVFGGGTKLTVL]Activatable antibody 15860-1204-SP34scFv(HvLv)-Fc fusionNucleotide sequence[spacer (SEQ ID NO: 507)][Activatable antibody 15860-1204-SP34scFv(HvLv)-Fcfusion without spacer (SEQ ID NO: 577)] (SEQ ID NO: 43)[CAAGGCCAGTCTGGCCAA][GTTTATTATTGCGGTGGGAATGAGAGTCTGTGCGGTGAGAGGAGGGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTGAAGTGCAGCTGGTGGAATCTGGGGGCGGACTGGTGCAGCCTAAGGGCAGCCTGAAGCTGAGCTGTGCCGCCAGCGGCTTCACCTTCAACACCTACGCCATGAACTGGGTGCGCCAGGCCCCTGGCAAAGGCCTGGAATGGGTGGCCCGGATCAGAAGCAAGTACAACAATTACGCCACCTACTACGCCGACAGCGTGAAGGACAGATTCACCATCAGCCGGGACGACAGCCAGAGCATCCTGTACCTGCAGATGAACAACCTGAAAACCGAGGACACCGCCATGTACTACTGCGTGCGGCACGGCAACTTCGGCAACAGCTATGTGTCTTGGTTTGCCTACTGGGGCCAGGGCACCCTCGTGACAGTGTCTGCCGGTGGTGGTGGTAGTGGTGGCGGTGGTTCAGGCGGTGGCGGTAGCCAGGCTGTCGTGACACAGGAAAGCGCCCTGACCACCAGCCCTGGCGAGACAGTGACCCTGACCTGTAGAAGCAGCACAGGCGCCGTGACCACAAGCAACTACGCCAATTGGGTGCAGGAAAAGCCCGACCACCTGTTCACCGGCCTGATCGGCGGCACCAACAAAAGGGCTCCAGGCGTGCCAGCCAGATTCAGCGGCTCCCTGATCGGAGATAAGGCCGCCCTGACAATCACTGGCGCCCAGACCGAGGACGAGGCCATCTACTTTTGCGCCCTGTGGTACAGCAACCTGTGGGTGTTCGGCGGAGGCACCAAGCTGACCGTGCTGGGCGGCTCCCTGGACCCTAAGTCATCTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACAACAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAA] Amino Acid sequence[spacer (SEQ ID NO: 87)][Activatable antibody 15860-1204-SP34scFv(HvLv)-Fcfusion without spacer (SEQ ID NO: 578)] (SEQ ID NO: 44)[QGQSGQ][VYYCGGNESLCGERRGSSGGSGGSGGLSGRSDNHGGGSEVQLVESGGGLVQPKGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSQSILYLQMNNLKTEDTAMYYCVRHGNFGNSYVSWFAYWGQGTLVTVSAGGGGSGGGGSGGGGSQAVVTQESALTTSPGETVTLTCRSSTGAVTTSNYANWVQEKPDHLFTGLIGGTNKRAPGVPARFSGSLIGDKAALTITGAQTEDEAIYFCALWYSNLWVFGGGTKLTVLGGSLDPKSSDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]15864-1204-SP34scFv(HvLv) Nucleotide sequence[spacer (SEQ ID NO: 507)][15864-1204-SP34scFv(HvLv) without spacer (SEQ IDNO: 579)] (SEQ ID NO: 45)[CAAGGCCAGTCTGGCCAA][TTTATGTGCCAGCAGCGGATGTGGGGGAATGAGTTTTGCCATCAGGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTGAAGTGCAGCTGGTGGAATCTGGGGGCGGACTGGTGCAGCCTAAGGGCAGCCTGAAGCTGAGCTGTGCCGCCAGCGGCTTCACCTTCAACACCTACGCCATGAACTGGGTGCGCCAGGCCCCTGGCAAAGGCCTGGAATGGGTGGCCCGGATCAGAAGCAAGTACAACAATTACGCCACCTACTACGCCGACAGCGTGAAGGACAGATTCACCATCAGCCGGGACGACAGCCAGAGCATCCTGTACCTGCAGATGAACAACCTGAAAACCGAGGACACCGCCATGTACTACTGCGTGCGGCACGGCAACTTCGGCAACAGCTATGTGTCTTGGTTTGCCTACTGGGGCCAGGGCACCCTCGTGACAGTGTCTGCCGGTGGTGGTGGTAGTGGTGGCGGTGGTTCAGGCGGTGGCGGTAGCCAGGCTGTCGTGACACAGGAAAGCGCCCTGACCACCAGCCCTGGCGAGACAGTGACCCTGACCTGTAGAAGCAGCACAGGCGCCGTGACCACAAGCAACTACGCCAATTGGGTGCAGGAAAAGCCCGACCACCTGTTCACCGGCCTGATCGGCGGCACCAACAAAAGGGCTCCAGGCGTGCCAGCCAGATTCAGCGGCTCCCTGATCGGAGATAAGGCCGCCCTGACAATCACTGGCGCCCAGACCGAGGACGAGGCCATCTACTTTTGCGCCCTGTGGTACAGCAACCTGTGGGTGTTCGGCGGAGGCACCAAGCTGACCGTGCTG] Amino Acid sequence[spacer (SEQ ID NO: 87)][15864-1204-SP34scFv(HvLv) without spacer (SEQ IDNO: 580)] (SEQ ID NO: 46)[QGQSGQ][FMCQQRMWGNEFCHQGSSGGSGGSGGLSGRSDNHGGGSEVQLVESGGGLVQPKGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSQSILYLQMNNLKTEDTAMYYCVRHGNFGNSYVSWFAYWGQGTLVTVSAGGGGSGGGGSGGGGSQAVVTQESALTTSPGETVTLTCRSSTGAVTTSNYANWVQEKPDHLFTGLIGGTNKRAPGVPARFSGSLIGDKAALTITGAQTEDEAIYFCALWYSNLWVFGGGTKLTVL]Activatable antibody 15864-1204-SP34scFv(HvLv)-Fc fusion[spacer (SEQ ID NO: 507)][Activatable antibody 15864-1204-SP34scFv(HvLv)-Fcfusion without spacer (SEQ ID NO: 581)] (SEQ ID NO: 47)[CAAGGCCAGTCTGGCCAA][TTTATGTGCCAGCAGCGGATGTGGGGGAATGAGTTTTGCCATCAGGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTGAAGTGCAGCTGGTGGAATCTGGGGGCGGACTGGTGCAGCCTAAGGGCAGCCTGAAGCTGAGCTGTGCCGCCAGCGGCTTCACCTTCAACACCTACGCCATGAACTGGGTGCGCCAGGCCCCTGGCAAAGGCCTGGAATGGGTGGCCCGGATCAGAAGCAAGTACAACAATTACGCCACCTACTACGCCGACAGCGTGAAGGACAGATTCACCATCAGCCGGGACGACAGCCAGAGCATCCTGTACCTGCAGATGAACAACCTGAAAACCGAGGACACCGCCATGTACTACTGCGTGCGGCACGGCAACTTCGGCAACAGCTATGTGTCTTGGTTTGCCTACTGGGGCCAGGGCACCCTCGTGACAGTGTCTGCCGGTGGTGGTGGTAGTGGTGGCGGTGGTTCAGGCGGTGGCGGTAGCCAGGCTGTCGTGACACAGGAAAGCGCCCTGACCACCAGCCCTGGCGAGACAGTGACCCTGACCTGTAGAAGCAGCACAGGCGCCGTGACCACAAGCAACTACGCCAATTGGGTGCAGGAAAAGCCCGACCACCTGTTCACCGGCCTGATCGGCGGCACCAACAAAAGGGCTCCAGGCGTGCCAGCCAGATTCAGCGGCTCCCTGATCGGAGATAAGGCCGCCCTGACAATCACTGGCGCCCAGACCGAGGACGAGGCCATCTACTTTTGCGCCCTGTGGTACAGCAACCTGTGGGTGTTCGGCGGAGGCACCAAGCTGACCGTGCTGGGCGGCTCCCTGGACCCTAAGTCATCTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACAACAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAA] Amino Acid sequence[spacer (SEQ ID NO: 87)][Activatable antibody 15864-1204-SP34scFv(HvLv)-Fcfusion without spacer (SEQ ID NO: 582)] (SEQ ID NO: 48)[QGQSGQ][FMCQQRMWGNEFCHQGSSGGSGGSGGLSGRSDNHGGGSEVQLVESGGGLVQPKGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSQSILYLQMNNLKTEDTAMYYCVRHGNFGNSYVSWFAYWGQGTLVTVSAGGGGSGGGGSGGGGSQAVVTQESALTTSPGETVTLTCRSSTGAVTTSNYANWVQEKPDHLFTGLIGGTNKRAPGVPARFSGSLIGDKAALTITGAQTEDEAIYFCALWYSNLWVFGGGTKLTVLGGSLDPKSSDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]15865-1204-SP34scFv(HvLv) Nucleotide sequence[spacer (SEQ ID NO: 507)][15865-1204-SP34scFv(HvLv) without spacer (SEQ IDNO: 583)] (SEQ ID NO: 49)[CAAGGCCAGTCTGGCCAA][ATGATGTATTGCGGTGGGAATGAGGTGTTGTGCGGGCCGCGGGTTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTGAAGTGCAGCTGGTGGAATCTGGGGGCGGACTGGTGCAGCCTAAGGGCAGCCTGAAGCTGAGCTGTGCCGCCAGCGGCTTCACCTTCAACACCTACGCCATGAACTGGGTGCGCCAGGCCCCTGGCAAAGGCCTGGAATGGGTGGCCCGGATCAGAAGCAAGTACAACAATTACGCCACCTACTACGCCGACAGCGTGAAGGACAGATTCACCATCAGCCGGGACGACAGCCAGAGCATCCTGTACCTGCAGATGAACAACCTGAAAACCGAGGACACCGCCATGTACTACTGCGTGCGGCACGGCAACTTCGGCAACAGCTATGTGTCTTGGTTTGCCTACTGGGGCCAGGGCACCCTCGTGACAGTGTCTGCCGGTGGTGGTGGTAGTGGTGGCGGTGGTTCAGGCGGTGGCGGTAGCCAGGCTGTCGTGACACAGGAAAGCGCCCTGACCACCAGCCCTGGCGAGACAGTGACCCTGACCTGTAGAAGCAGCACAGGCGCCGTGACCACAAGCAACTACGCCAATTGGGTGCAGGAAAAGCCCGACCACCTGTTCACCGGCCTGATCGGCGGCACCAACAAAAGGGCTCCAGGCGTGCCAGCCAGATTCAGCGGCTCCCTGATCGGAGATAAGGCCGCCCTGACAATCACTGGCGCCCAGACCGAGGACGAGGCCATCTACTTTTGCGCCCTGTGGTACAGCAACCTGTGGGTGTTCGGCGGAGGCACCAAGCTGACCGTGCTG] Amino Acid sequence[spacer (SEQ ID NO: 87)][15865-1204-SP34scFv(HvLv) without spacer (SEQ IDNO: 584)] (SEQ ID NO: 50)[QGQSGQ][MMYCGGNEVLCGPRVGSSGGSGGSGGLSGRSDNHGGGSEVQLVESGGGLVQPKGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSQSILYLQMNNLKTEDTAMYYCVRHGNFGNSYVSWFAYWGQGTLVTVSAGGGGSGGGGSGGGGSQAVVTQESALTTSPGETVTLTCRSSTGAVTTSNYANWVQEKPDHLFTGLIGGTNKRAPGVPARFSGSLIGDKAALTITGAQTEDEAIYFCALWYSNLWVFGGGTKLTVL]Activatable antibody 15865-1204-SP34 scFv(HvLv)-Fc fusionNucleotide sequence[spacer (SEQ ID NO: 507)][Activatable antibody 15865-1204-SP34scFv(HvLv)-Fcfusion without spacer (SEQ ID NO: 585)] (SEQ ID NO: 51)[CAAGGCCAGTCTGGCCAA][ATGATGTATTGCGGTGGGAATGAGGTGTTGTGCGGGCCGCGGGTTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTGAAGTGCAGCTGGTGGAATCTGGGGGCGGACTGGTGCAGCCTAAGGGCAGCCTGAAGCTGAGCTGTGCCGCCAGCGGCTTCACCTTCAACACCTACGCCATGAACTGGGTGCGCCAGGCCCCTGGCAAAGGCCTGGAATGGGTGGCCCGGATCAGAAGCAAGTACAACAATTACGCCACCTACTACGCCGACAGCGTGAAGGACAGATTCACCATCAGCCGGGACGACAGCCAGAGCATCCTGTACCTGCAGATGAACAACCTGAAAACCGAGGACACCGCCATGTACTACTGCGTGCGGCACGGCAACTTCGGCAACAGCTATGTGTCTTGGTTTGCCTACTGGGGCCAGGGCACCCTCGTGACAGTGTCTGCCGGTGGTGGTGGTAGTGGTGGCGGTGGTTCAGGCGGTGGCGGTAGCCAGGCTGTCGTGACACAGGAAAGCGCCCTGACCACCAGCCCTGGCGAGACAGTGACCCTGACCTGTAGAAGCAGCACAGGCGCCGTGACCACAAGCAACTACGCCAATTGGGTGCAGGAAAAGCCCGACCACCTGTTCACCGGCCTGATCGGCGGCACCAACAAAAGGGCTCCAGGCGTGCCAGCCAGATTCAGCGGCTCCCTGATCGGAGATAAGGCCGCCCTGACAATCACTGGCGCCCAGACCGAGGACGAGGCCATCTACTTTTGCGCCCTGTGGTACAGCAACCTGTGGGTGTTCGGCGGAGGCACCAAGCTGACCGTGCTGGGCGGCTCCCTGGACCCTAAGTCATCTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACAACAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAA] Amino Acid sequence[spacer (SEQ ID NO: 87)][Activatable antibody 15865-1204-SP34scFv(HvLv)-Fcfusion without spacer (SEQ ID NO: 586)] (SEQ ID NO: 52)[QGQSGQ][MMYCGGNEVLCGPRVGSSGGSGGSGGLSGRSDNHGGGSEVQLVESGGGLVQPKGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSQSILYLQMNNLKTEDTAMYYCVRHGNFGNSYVSWFAYWGQGTLVTVSAGGGGSGGGGSGGGGSQAVVTQESALTTSPGETVTLTCRSSTGAVTTSNYANWVQEKPDHLFTGLIGGTNKRAPGVPARFSGSLIGDKAALTITGAQTEDEAIYFCALWYSNLWVFGGGTKLTVLGGSLDPKSSDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]

Example 3 In Vitro Characterization of Activatable Anti-CD3ε Antibodies

This Example describes the ability of a masking moiety of the disclosureto reduce the ability of activatable anti-CD3ε antibodies of thedisclosure comprising such a masking moiety to bind to CD3ε.

To determine if the masking peptides described in the previous examplescould inhibit binding in the SP34 scFv-Fc context, a flowcytometry-based binding assay was performed. Jurkat T cells (Clone E6-1,ATCC, TIB-152) were cultured in RPMI-1640+glutamax (Life Technologies,Catalog 72400-120), 10% Heat Inactivated-Fetal Bovine Serum (HI-FBS,Life Technologies, Catalog 10438-026), 100 U/ml penicillin, and 100μg/ml streptomycin (Life Technologies, Catalog 15140-122) according toATCC guidelines. Cells were harvested by centrifugation (200×g, 4° C., 5min) and re-suspended in PBS supplemented with 2% HI-FBS (FACS Buffer).About 180,000 Jurkats per well were transferred to a 96-well U-bottomplate, harvested, and re-suspended in 50 μL of primary antibody. Thestarting concentration of activatable antibody was 1 μM followed by3-fold serial dilutions for a total of 12 concentrations. That followingactivatable antibodies were tested; 15003-1204-SP34scFv(LvHv)-Fc fusion;15860-1204-SP34scFv(LvHv)-Fc fusion; 15864-1204-SP34scFv(LvHv)-Fcfusion; and 15865-1204-SP34scFv(LvHv)-Fc fusion; as was antibodySP34scFv(LvHv)-Fc.

Cells were incubated at 4° C. with shaking for about 1 hour, harvested,and washed with 3×200 μL of FACS Buffer. Jurkats were resuspended in 50μl Alexa Fluor 488 conjugated anti-Human IgG Fc (1:100 dilution, JacksonImmunoResearch, Product 109-546-098) and incubated at 4° C. with shakingfor about 30 min. Cells were harvested, washed with 3×200 μL of FACSBuffer, and resuspended in a final volume of 120 μL of FACS Buffer.Samples were collected on a BD Accuri C6 (BD Biosciences) and the medianfluorescence intensity (MFI) of viable cells was calculated using FlowJoV10 (Treestar). EC₅₀ values were calculated in GraphPad Prism 6 by curvefitting the data to log(agonist) vs. response (three parameters).

FIGS. 3A-3D demonstrate that incorporation of the masking peptides intothe SP34 scFv-Fc context to produce activatable antibodies shifted theEC₅₀ values for CD3ε binding from single digit to triple digit nM.

Example 4 Bispecific Activatable Antibodies

This Example describes the production, expression and characterizationof bispecific activatable antibodies of the disclosure.

Vector Construction:

The heavy chain and light chain were cloned separately into a mammalianexpression vector using standard molecular biology techniques. Briefly,DNA fragments encoding the region of interest were amplified withprimers binding to the terminal ends. Overlapping fragments werecombined and amplified with flanking primers as needed to build theentire desired region. DNA fragments were subsequently cloned intoexpression vector using a commercially available homologousrecombination kit (MCLabs, South San Francisco, Calif.). The in-housemammalian expression vector is a modified version of cDNA3.1(+) fromInvitrogen with selection marker of G418 or hygromycin. Mutations wereintroduced using the QuikChange Kit (Agilent, Santa Clara, Calif.).

Bispecific antibodies and bispecific activatable antibodies wereexpressed in mammalian cells using standard transfection kit (LifeTechnologies, Grand Island, N.Y.). Briefly, 293 cells were transfectedwith nucleic acids using a lipid based system, following manufacturesrecommended protocol. Bispecific activatable antibodies and bispecificantibodies were purified from cell-free supernatant using Protein Abeads (GE, Piscataway, N.J.) and concentrated using standard bufferexchange columns (Millipore, Temecula, Calif.).

Table 11 below describes the heavy chains and light chains for variousbispecific antibodies and bispecific activatable antibodies of thedisclosure.

TABLE 11 Bispecific antibodies and bispecific activatable antibodiesMolecule Name Molecule Heavy Chain Light Chain CI005 C225v5N297Q-CD3-pLW019: HC OPP007: LC C225 H-N C225v5N297Q- CD3HvLv-H-N CI007 3954-1204-pLW019: HC OPP022: LC C225 3954-1204 C225v5N297Q-CD3- C225v5N297Q- H-NCD3HvLv-H-N CI009 C225v5N297Q-15865- pLW023: HC OPP007: LC C225CD3LvHv-H-N C225v5N297Q-15865- CD3LvHv-H-N CI010 3954-1204- pLW022: HCOPP022: LC C225 3954-1204 C225v5N297Q-15003- C225v5N297Q-15003-CD3LvHv-H-N CD3LvHv-H-N CI011 3954-1204- pLW023: HC OPP022: LC C2253954-1204 C225v5N297Q-15865- C225v5N297Q-15865- CD3LvHv-H-N CD3LvHv-H-NCI012 C225v5N297Q pLW006: HC OPP021: LC C225 C225v5N297Q CI015C225v5N297Q- pLW057: HC OPP021: LC C225 CD3LvHv-H-N C225v5N297Q-CD3LvHv-H-N CI016 3954-1204- pLW057: HC OPP022: LC C225 3954-1204C225v5N297Q- C225v5N297Q- CD3LvHv-H-N CD3LvHv-H-N CI021C225v5N297Q-15003- pLW047: HC OPP021: LC C225 CD3HvLv-H-NC225v5N297Q-15003- CD3HvLv-H-N CI022 C225v5N297Q-15865- pLW048: HCOPP021: LC C225 CD3HvLv-H-N C225v5N297Q-15865- CD3HvLv-H-N CI0233954-1204- pLW047: HC OPP022: LC C225 3954-1204 C225v5N297Q-15003-C225v5N297Q-15003- CD3HvLv-H-N CD3HvLv-H-N CI024 3954-1204- pLW048: HCOPP022: LC C225 3954-1204 C225v5N297Q-15865- C225v5N297Q-15865-CD3HvLv-H-N CD3HvLv-H-N CI025 IL6RN297Q pLW078: HC pLW077: LC IL6RIL6RN297Q CI026 IL6RN297Q- pLW083: HC pLW077: LC IL6R CD3LvHv-H-NIL6RN297Q-CD3LvHv- H-N CI027 IL6RN297Q- pLW085: HC pLW077: LC IL6R15865_1204- IL6RN297Q-15865_1204- CD3LvHv-H-N CD3LvHv-H-N CI028 IL6R4792 Nsub pLW085: HC pLW079: LC IL6R 4792 Nsub N297Q-15865_1204-IL6RN297Q-15865_1204- CD3LvHv-H-N CD3LvHv-H-N CI029 4792-1204- pLW085:HC pLW080: LC IL6R 4792 1204 IL6RN297Q- IL6RN297Q-15865_1204- (C040)15865_1204- CD3LvHv-H-N CD3LvHv-H-N CI030 IL6RN297Q- pLW087: HC pLW077:LC IL6R 15865_Nsub- IL6RN297Q- CD3LvHv-H-N 15865_Nsub-CD3LvHv- H-N CI031IL6R 4792 pLW087: HC pLW079: LC IL6R 4792 Nsub NsubN297Q- IL6RN297Q-15865_Nsub- 15865_Nsub-CD3LvHv- CD3LvHv-H-N H-N CI032 4792-1204- pLW087:HC pLW080: LC IL6R 4792 1204 IL6RN297Q- IL6RN297Q- 15865_Nsub-15865_Nsub-CD3LvHv- CD3LvHv-H-N H-N CI036 4792-1204- pLW083: HC pLW080:LC IL6R 4792 1204 IL6RN297Q- IL6RN297Q-CD3LvHv- CD3LvHv-H-N H-N CI0393954-2001-C225v5 pLW101: HC LC C225-3954-2001 N297Q-15865-2001-C225v5N297Q-15865- CD3LvHv-H-N 2001-CD3LvHv-H-N CI040 3954- pLW101: HCLC C225-3954-2001 2001C225v5N297Q- C225v5N297Q-15865- 15865-2001-2001-CD3LvHv-H-N CD3LvHv-H-N CI048 activated bispecific pLW023: HCOPP022: LC C225 3954-1204 activatable antibody C225v5N297Q-15865-C225N297Q- CD3LvHv-H-N *CD3LvHv-H-N CI049 3954-1204- pLW121: HC OPP022:LC C225 3954-1204 C225v5N297Q-MC05- C225v5N297Q-MC05- 2001-mCD3-H-N2001-mCD3-H-N CI050 3954-1204- pLW122: HC OPP022: LC C225 3954-1204C225v5N297Q-MC06- C225v5N297Q-MC06- 2001-mCD3-H-N 2001-mCD3-H-N CI0513954-1204- pLW123: HC OPP022: LC C225 3954-1204 C225v5N297Q-MC07-C225v5N297Q-MC07- 2001-mCD3-H-N 2001-mCD3-H-N CI052 C225v5N297Q-mCD3-pLW100: HC OPP021: LC C225 H-N C225v5N297Q-mCD3-H-N CI053 3954-1204-pLW117: HC OPP022: LC C225 3954-1204 C225v5N297Q-MC01- C225v5N297Q-MC01-2001-mCD3-H-N 2001-mCD3-H-N CI054 3954-1204- pLW118: HC OPP022: LC C2253954-1204 C225v5N297Q-MC02- C225v5N297Q-MC02- 2001-mCD3-H-N2001-mCD3-H-N CI055 3954-1204- pLW119: HC OPP022: LC C225 3954-1204C225v5N297Q-MC03- C225v5N297Q-MC03- 2001-mCD3-H-N 2001-mCD3-H-N CI0563954-1204- pLW120: HC OPP022: LC C225 3954-1204 C225v5N297Q-MC04-C225v5N297Q-MC04- 2001-mCD3-H-N 2001-mCD3-H-N CI057 3954-1204- pLW124:HC OPP022: LC C225 3954-1204 C225v5N297Q-MC08- C225v5N297Q-MC08-2001-mCD3-H-N 2001-mCD3-H-N CI058 3954-1204- pLW125: HC OPP022: LC C2253954-1204 C225v5N297Q-MC09- C225v5N297Q-MC09- 2001-mCD3-H-N2001-mCD3-H-N CI059 BiTE BiTE

Table 12A provide the primers used for each construct, and Table 12Bprovides the sequences of the primers used.

TABLE 12A Primers Construct Name Primers Used in Construction BiTEoLW017, oLW018 pLW006 oLW006, oLW007 pLW019 oLW048, oLW049, oLW050,oLW051 pLW022 oLW048, oLW061, oLW062, oLW063 pLW023 oLW048, oLW061,oLW062, oLW063 pLW047 oLW048, oLW049, oLW051, oLW098, oLW101, oLW102pLW048 oLW048, oLW049, oLW051, oLW098, oLW101, oLW103 pLW057 oLW048,oLW056, oLW063, oLW109 pLW077 oLW048, oLW056, oLW115, oLW116 pLW078oLW048, oLW049, oLW117, oLW118, oLW119, oLW120 pLW080 oLW048, oLW121pLW083 oLW048, oLW123, oLW124, oLW125 pLW085 oLW048, oLW123, oLW124,oLW125 pLW101 oLW048, oLW151, oLW062, oLW063 pLW117 oLW048, oLW049,oLW178 pLW118 oLW048, oLW049, oLW178 pLW119 oLW048, oLW049, oLW178pLW120 oLW048, oLW049, oLW178 pLW121 oLW048, oLW049, oLW178 pLW122oLW048, oLW049, oLW178 pLW123 oLW048, oLW049, oLW178 pLW124 oLW048,oLW049, oLW178 pLW125 oLW048, oLW049, oLW178

TABLE 12B  Primer Sequences Primer SEQ ID Name Primer Sequence NO:oLW001 CATACACTGGTATCAGCAAAGAACACAGGGTTCTCC 412 AAGGC oLW002GCCTTGGAGAACCCTGTGTTCTTTGCTGATACCAGT 413 GTATG oLW003TTTAAAATGAACAGTCTGCAATCTCAGGACACAGCC 414 ATATATTACTGTGCC oLW004GGCACAGTAATATATGGCTGTGTCCTGAGATTGCAG 415 ACTGTTCATTTTAAA oLW006CGGGAGGAGCAGTACCAGAGCACGTACCGTGTG 416 oLW007CACACGGTACGTGCTCTGGTACTGCTCCTCCCG 417 oLW017 CACCCCATTGACGTCAATGGGAG 418oLW018 GAGGGGCAAACAACAGATGGCTG 419 oLW048TGAACCGTCAGATCACTAGAAGCTTTATTGC 420 oLW049 TGGATGTGCACCAGGTGTGACAGC 421oLW050 CTGGCTGTCACACCTGGTGCACATCCAGAGGTGCAG 422 CTGGTCGAGTCTGG oLW051AGCTGCACCTGGGATCCACCACCTCCTAGGACAGTC 423 AGTTTGGTTCCTCCACC oLW056ACATCTAGCACCACGCAGCCATAGTAGC 424 oLW061GTGACCACGGTCTGAGAACCGCCGCCATGATTATCG 425 G oLW062GGCGGCGGTTCTCAGACCGTGGTCACACAGGAGCC 426 oLW063CTCTGTTTCAGCTGCACCTGGGATCCACCACCTCCT 427 GAGGAGACGGTGACCAGTGTCCC oLW098AGAACCGCCGCCATGATTATCG 428 oLW101 CGTTCCGATAATCATGGCGGCGGTTCTGAGGTGCAG429 CTGGTCGAGTCTGG oLW102 CTGGCTGTCACACCTGGTGCACATCCACAAGGCCAG 430TCTGGCCAAGGTTATC oLW103 CTGGCTGTCACACCTGGTGCACATCCACAAGGCCAG 431TCTGGCCAAATGATG oLW109 CTCCTGCTACTATGGCTGCGTGGTGCTAGATGTCAG 432ACCGTGGTCACACAGGAGCC oLW115 CTCCTGCTACTATGGCTGCGTGGTGCTAGATGTGAC 433ATCCAGATGACTCAGTCTCCTAGCTCC oLW116 GAAGACAGATGGTGCAGCCACCG 434 oLW117CTGGCTGTCACACCTGGTGCACATCCACAGGTGCAG 435 CTGCAGGAGTCCG oLW118GCTAGCACCAAGGGCCCATCG 436 oLW119 GGAAGACCGATGGGCCCTTGG 437 oLW120GCCCTCTAGACTCGAGCGGCCGCTCATTTACCCGGA 438 GACAGGGAGAG oLW121GCAGGACCCATACTGGCCAGACTGGCCTTGACATCT 439 AGCACCACGCAGCCATAGTAGC oLW123GGAGGTGGTGGATCCCAGGTGCAGCTGCAGGAGTCC 440 G oLW124CAGCTGCACCTGGGATCCACCACCTCCTGAGGAGAC 441 G oLW125 ACCGATGGGCCCTTGGTGCTAG442 oLW151 GACCACGGTCTGAGAACCGCCG 443 oLW178 CACCTGGTGCACATCCACAAGGC 444

The sequences for the bispecific antibodies and bispecific activatableantibodies are shown below. In some embodiments, the activatableantibody also includes a spacer sequence. In some embodiments, thespacer is joined directly to the MM of the activatable antibody. In someembodiments, the spacer is joined directly to the MM of the activatableantibody in the structural arrangement from N-terminus to C-terminus ofspacer-MM-CM-AB. In some embodiments, the spacer joined directly to theN-terminus of MM of the activatable antibody is selected from the groupconsisting of QGQSGQG (SEQ ID NO: 407), QGQSGQ (SEQ ID NO: 87), QGQSG(SEQ ID NO: 408), QGQS (SEQ ID NO: 409), QGQ (SEQ ID NO: 410), QG (SEQID NO: 411), and Q. In some embodiments, the spacer includes at leastthe amino acid sequence QGQSGQG (SEQ ID NO: 412). In some embodiments,the spacer includes at least the amino acid sequence QGQSGQG (SEQ ID NO:407). In some embodiments, the spacer includes at least the amino acidsequence QGQSGQ (SEQ ID NO: 87). In some embodiments, the spacerincludes at least the amino acid sequence QGQSG (SEQ ID NO: 408). Insome embodiments, the spacer includes at least the amino acid sequenceQGQS (SEQ ID NO: 409), In some embodiments, the spacer includes at leastthe amino acid sequence QGQ (SEQ ID NO: 410). In some embodiments, thespacer includes at least the amino acid sequence QG (SEQ ID NO: 411). Insome embodiments, the spacer includes at least the amino acid residue Q.

While some of the sequences shown below include the spacer sequence ofSEQ ID NO: 362, those of ordinary skill in the art appreciate that themultispecific activatable antibodies of the disclosure can include anysuitable spacer sequence, such as, for example, a spacer sequenceselected from the group consisting of QGQSGQG (SEQ ID NO: 407), QGQSGQ(SEQ ID NO: 87), QGQSG (SEQ ID NO: 408), QGQS (SEQ ID NO: 409), QGQ (SEQID NO: 410), QG (SEQ ID NO: 411), and Q.

CI005: C225v5N297Q-CD3-H-N pLW019: HC C225v5N297Q-CD3HvLv-H-NNucleotide Sequence (SEQ ID NO: 445)GAGGTGCAGCTGGTCGAGTCTGGAGGAGGATTGGTGCAGCCTGGAGGGTCATTGAAACTCTCATGTGCAGCCTCTGGATTCACCTTCAATAAGTACGCCATGAACTGGGTCCGCCAGGCTCCAGGAAAGGGTTTGGAATGGGTTGCTCGCATAAGAAGTAAATATAATAATTATGCAACATATTATGCCGATTCAGTGAAAGACAGGTTCACCATCTCCAGAGATGATTCAAAAAACACTGCCTATCTACAAATGAACAACTTGAAAACTGAGGACACTGCCGTGTACTACTGTGTGAGACATGGGAACTTCGGTAATAGCTACATATCCTACTGGGCTTACTGGGGCCAAGGGACTCTGGTCACCGTCTCCTCAGGTGGTGGTGGTTCTGGCGGCGGCGGCTCCGGTGGTGGTGGTTCTCAGACTGTTGTGACTCAGGAACCTTCACTCACCGTATCACCTGGTGGAACAGTCACACTCACTTGTGGCTCCTCGACTGGGGCTGTTACATCTGGCTACTACCCAAACTGGGTCCAACAAAAACCAGGTCAGGCACCCCGTGGTCTAATAGGTGGGACTAAGTTCCTCGCCCCCGGTACTCCTGCCAGATTCTCAGGCTCCCTGCTTGGAGGCAAGGCTGCCCTCACCCTCTCAGGGGTACAGCCAGAGGATGAGGCAGAATATTACTGTGCTCTATGGTACAGCAACCGCTGGGTGTTCGGTGGAGGAACCAAACTGACTGTCCTAGGAGGTGGTGGATCCCAGGTGCAGCTGAAACAGAGCGGCCCGGGCCTGGTGCAGCCGAGCCAGAGCCTGAGCATTACCTGCACCGTGAGCGGCTTTAGCCTGACCAACTATGGCGTGCATTGGGTGCGCCAGAGCCCGGGCAAAGGCCTGGAATGGCTGGGCGTGATTTGGAGCGGCGGCAACACCGATTATAACACCCCGTTTACCAGCCGCCTGAGCATTAACAAAGATAACAGCAAAAGCCAGGTGTTTTTTAAAATGAACAGCCTGCAAAGCCAGGATACCGCGATTTATTATTGCGCGCGCGCGCTGACCTATTATGATTATGAATTTGCGTATTGGGGCCAGGGCACCCTGGTGACCGTGAGCGCGGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA  Amino Acid Sequence (SEQ ID NO: 446)EVQLVESGGGLVQPGGSLKLSCAASGFTFNKYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKNTAYLQMNNLKTEDTAVYYCVRHGNFGNSYISYWAYWGQGTLVTVSSGGGGSGGGGSGGGGSQTVVTQEPSLTVSPGGTVTLTCGSSTGAVTSGYYPNWVQQKPGQAPRGLIGGTKFLAPGTPARFSGSLLGGKAALTLSGVQPEDEAEYYCALWYSNRWVFGGGTKLTVLGGGGSQVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSQDTAIYYCARALTYYDYEFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK*  OPP007: LC C225 IL2ss Nucleotide Sequence (SEQ ID NO: 447)GATATCTTGCTGACCCAGAGCCCGGTGATTCTGAGCGTGAGCCCGGGCGAACGTGTGAGCTTTAGCTGCCGCGCGAGCCAGAGCATTGGCACCAACATTCATTGGTATCAGCAGCGCACCAACGGCAGCCCGCGCCTGCTGATTAAATATGCGAGCGAAAGCATTAGCGGCATTCCGAGCCGCTTTAGCGGCAGCGGCAGCGGCACCGATTTTACCCTGAGCATTAACAGCGTGGAAAGCGAAGATATTGCGGATTATTATTGCCAGCAGAACAACAACTGGCCGACCACCTTTGGCGCGGGCACCAAACTGGAACTGAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG  Amino Acid Sequence (SEQ ID NO: 448)DILLTQSPVILSVSPGERVSFSCRASQSIGTNIHWYQQRTNGSPRLLIKYASESISGIPSRFSGSGSGTDFTLSINSVESEDIADYYCQQNNNWPTTFGAGTKLELKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC* CI007: 3954-1204-C225v5N297Q-CD3-H-N pLW019: HC C225v5N297Q-CD3HvLv-H-NNucleotide Sequence (SEQ ID NO: 445)GAGGTGCAGCTGGTCGAGTCTGGAGGAGGATTGGTGCAGCCTGGAGGGTCATTGAAACTCTCATGTGCAGCCTCTGGATTCACCTTCAATAAGTACGCCATGAACTGGGTCCGCCAGGCTCCAGGAAAGGGTTTGGAATGGGTTGCTCGCATAAGAAGTAAATATAATAATTATGCAACATATTATGCCGATTCAGTGAAAGACAGGTTCACCATCTCCAGAGATGATTCAAAAAACACTGCCTATCTACAAATGAACAACTTGAAAACTGAGGACACTGCCGTGTACTACTGTGTGAGACATGGGAACTTCGGTAATAGCTACATATCCTACTGGGCTTACTGGGGCCAAGGGACTCTGGTCACCGTCTCCTCAGGTGGTGGTGGTTCTGGCGGCGGCGGCTCCGGTGGTGGTGGTTCTCAGACTGTTGTGACTCAGGAACCTTCACTCACCGTATCACCTGGTGGAACAGTCACACTCACTTGTGGCTCCTCGACTGGGGCTGTTACATCTGGCTACTACCCAAACTGGGTCCAACAAAAACCAGGTCAGGCACCCCGTGGTCTAATAGGTGGGACTAAGTTCCTCGCCCCCGGTACTCCTGCCAGATTCTCAGGCTCCCTGCTTGGAGGCAAGGCTGCCCTCACCCTCTCAGGGGTACAGCCAGAGGATGAGGCAGAATATTACTGTGCTCTATGGTACAGCAACCGCTGGGTGTTCGGTGGAGGAACCAAACTGACTGTCCTAGGAGGTGGTGGATCCCAGGTGCAGCTGAAACAGAGCGGCCCGGGCCTGGTGCAGCCGAGCCAGAGCCTGAGCATTACCTGCACCGTGAGCGGCTTTAGCCTGACCAACTATGGCGTGCATTGGGTGCGCCAGAGCCCGGGCAAAGGCCTGGAATGGCTGGGCGTGATTTGGAGCGGCGGCAACACCGATTATAACACCCCGTTTACCAGCCGCCTGAGCATTAACAAAGATAACAGCAAAAGCCAGGTGTTTTTTAAAATGAACAGCCTGCAAAGCCAGGATACCGCGATTTATTATTGCGCGCGCGCGCTGACCTATTATGATTATGAATTTGCGTATTGGGGCCAGGGCACCCTGGTGACCGTGAGCGCGGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA  Amino Acid Sequence (SEQ ID NO: 446)EVQLVESGGGLVQPGGSLKLSCAASGFTFNKYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKNTAYLQMNNLKTEDTAVYYCVRHGNFGNSYISYWAYWGQGTLVTVSSGGGGSGGGGSGGGGSQTVVTQEPSLTVSPGGTVTLTCGSSTGAVTSGYYPNWVQQKPGQAPRGLIGGTKFLAPGTPARFSGSLLGGKAALTLSGVQPEDEAEYYCALWYSNRWVFGGGTKLTVLGGGGSQVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSQDTAIYYCARALTYYDYEFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK*  OPP022: LC C225 3954-1204 Nucleotide Sequence[spacer (SEQ ID NO: 507)][OPP022 without spacer (SEQ ID NO: 509)](SEQ ID NO: 449)[CAAGGCCAGTCTGGCCAG][TGCATCTCACCTCGTGGTTGTCCGGACGGCCCATACGTCATGTACGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGATCCGGTCTGAGCGGCCGTTCCGATAATCATGGCAGTAGCGGTACCCAGATCTTGCTGACCCAGAGCCCGGTGATTCTGAGCGTGAGCCCGGGCGAACGTGTGAGCTTTAGCTGCCGCGCGAGCCAGAGCATTGGCACCAACATTCATTGGTATCAGCAGCGCACCAACGGCAGCCCGCGCCTGCTGATTAAATATGCGAGCGAAAGCATTAGCGGCATTCCGAGCCGCTTTAGCGGCAGCGGCAGCGGCACCGATTTTACCCTGAGCATTAACAGCGTGGAAAGCGAAGATATTGCGGATTATTATTGCCAGCAGAACAACAACTGGCCGACCACCTTTGGCGCGGGCACCAAACTGGAACTGAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG] Amino Acid Sequence[spacer (SEQ ID NO: 87)][OPP022 without spacer (SEQ ID NO: 508)](SEQ ID NO: 450)[QGQSGQ][CISPRGCPDGPYVMYGSSGGSGGSGGSGLSGRSDNHGSSGTQILLTQSPVILSVSPGERVSFSCRASQSIGTNIHWYQQRTNGSPRLLIKYASESISGIPSRFSGSGSGTDFTLSINSVESEDIADYYCQQNNNWPTTFGAGTKLELKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC]* CI009: C225v5N297Q-15865-CD3LvHv-H-NpLW023: C225v5N297Q-15865-CD3LvHv-H-N Nucleotide Sequence[spacer (SEQ ID NO: 507)][pLW023 without spacer (SEQ ID NO: 511)](SEQ ID NO: 451)[CAAGGCCAGTCTGGCCAA][ATGATGTATTGCGGTGGGAATGAGGTGTTGTGCGGGCCGCGGGTTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTCAGACCGTGGTCACACAGGAGCCCTCACTGACAGTGAGCCCTGGCGGGACCGTCACACTGACTTGTCGCAGTTCAACTGGCGCCGTGACTACCAGCAATTACGCTAACTGGGTCCAGCAGAAACCAGGACAGGCACCACGAGGACTGATCGGAGGAACTAATAAGAGAGCACCAGGAACCCCTGCAAGGTTCTCCGGATCTCTGCTGGGGGGAAAAGCCGCTCTGACACTGAGCGGCGTGCAGCCTGAGGACGAAGCTGAGTACTATTGCGCACTGTGGTACTCCAACCTGTGGGTGTTTGGCGGGGGAACTAAGCTGACCGTCCTGGGAGGAGGAGGAAGCGGAGGAGGAGGGAGCGGAGGAGGAGGATCCGAAGTGCAGCTGGTCGAGAGCGGAGGAGGACTGGTGCAGCCAGGAGGATCCCTGAAGCTGTCTTGTGCAGCCAGTGGCTTCACCTTCAACACTTACGCAATGAACTGGGTGCGGCAGGCACCTGGGAAGGGACTGGAATGGGTCGCCCGGATCAGATCTAAATACAATAACTATGCCACCTACTATGCTGACAGTGTGAAGGATAGGTTCACCATTTCACGCGACGATAGCAAAAACACAGCTTATCTGCAGATGAATAACCTGAAGACCGAGGATACAGCAGTGTACTATTGCGTCAGACACGGCAATTTCGGGAACTCTTACGTGAGTTGGTTTGCCTATTGGGGACAGGGGACACTGGTCACCGTCTCCTCAGGAGGTGGTGGATCCCAGGTGCAGCTGAAACAGAGCGGCCCGGGCCTGGTGCAGCCGAGCCAGAGCCTGAGCATTACCTGCACCGTGAGCGGCTTTAGCCTGACCAACTATGGCGTGCATTGGGTGCGCCAGAGCCCGGGCAAAGGCCTGGAATGGCTGGGCGTGATTTGGAGCGGCGGCAACACCGATTATAACACCCCGTTTACCAGCCGCCTGAGCATTAACAAAGATAACAGCAAAAGCCAGGTGTTTTTTAAAATGAACAGCCTGCAAAGCCAGGATACCGCGATTTATTATTGCGCGCGCGCGCTGACCTATTATGATTATGAATTTGCGTATTGGGGCCAGGGCACCCTGGTGACCGTGAGCGCGGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA] Amino Acid Sequence[spacer (SEQ ID NO: 87)][pLW023 without spacer (SEQ ID NO: 512)](SEQ ID NO: 452)[QGQSGQ][MMYCGGNEVLCGPRVGSSGGSGGSGGLSGRSDNHGGGSQTVVTQEPSLTVSPGGTVTLTCRSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGTPARFSGSLLGGKAALTLSGVQPEDEAEYYCALWYSNLWVFGGGTKLTVLGGGGSGGGGSGGGGSEVQLVESGGGLVQPGGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKNTAYLQMNNLKTEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSSGGGGSQVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSQDTAIYYCARALTYYDYEFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]*OPP007: LC C225 IL2ss Nucleotide Sequence (SEQ ID NO: 447)GATATCTTGCTGACCCAGAGCCCGGTGATTCTGAGCGTGAGCCCGGGCGAACGTGTGAGCTTTAGCTGCCGCGCGAGCCAGAGCATTGGCACCAACATTCATTGGTATCAGCAGCGCACCAACGGCAGCCCGCGCCTGCTGATTAAATATGCGAGCGAAAGCATTAGCGGCATTCCGAGCCGCTTTAGCGGCAGCGGCAGCGGCACCGATTTTACCCTGAGCATTAACAGCGTGGAAAGCGAAGATATTGCGGATTATTATTGCCAGCAGAACAACAACTGGCCGACCACCTTTGGCGCGGGCACCAAACTGGAACTGAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG  Amino Acid Sequence (SEQ ID NO: 448)DILLTQSPVILSVSPGERVSFSCRASQSIGTNIHWYQQRTNGSPRLLIKYASESISGIPSRFSGSGSGTDFTLSINSVESEDIADYYCQQNNNWPTTFGAGTKLELKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC* CI010: 3954-1204-C225v5N297Q-15003-CD3LvHv-H-NpLW022: C225v5N297Q-15865-CD3LvHv-H-N Nucleotide Sequence[spacer (SEQ ID NO: 507)][pLW022 without spacer (SEQ ID NO: 895)](SEQ ID NO: 451)[CAAGGCCAGTCTGGCCAA][GGTTATCGGTGGGGTTGCGAGTGGAATTGCGGTGGGATTACTACTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTCAGACCGTGGTCACACAGGAGCCCTCACTGACAGTGAGCCCTGGCGGGACCGTCACACTGACTTGTCGCAGTTCAACTGGCGCCGTGACTACCAGCAATTACGCTAACTGGGTCCAGCAGAAACCAGGACAGGCACCACGAGGACTGATCGGAGGAACTAATAAGAGAGCACCAGGAACCCCTGCAAGGTTCTCCGGATCTCTGCTGGGGGGAAAAGCCGCTCTGACACTGAGCGGCGTGCAGCCTGAGGACGAAGCTGAGTACTATTGCGCACTGTGGTACTCCAACCTGTGGGTGTTTGGCGGGGGAACTAAGCTGACCGTCCTGGGAGGAGGAGGAAGCGGAGGAGGAGGGAGCGGAGGAGGAGGATCCGAAGTGCAGCTGGTCGAGAGCGGAGGAGGACTGGTGCAGCCAGGAGGATCCCTGAAGCTGTCTTGTGCAGCCAGTGGCTTCACCTTCAACACTTACGCAATGAACTGGGTGCGGCAGGCACCTGGGAAGGGACTGGAATGGGTCGCCCGGATCAGATCTAAATACAATAACTATGCCACCTACTATGCTGACAGTGTGAAGGATAGGTTCACCATTTCACGCGACGATAGCAAAAACACAGCTTATCTGCAGATGAATAACCTGAAGACCGAGGATACAGCAGTGTACTATTGCGTCAGACACGGCAATTTCGGGAACTCTTACGTGAGTTGGTTTGCCTATTGGGGACAGGGGACACTGGTCACCGTCTCCTCAGGAGGTGGTGGATCCCAGGTGCAGCTGAAACAGAGCGGCCCGGGCCTGGTGCAGCCGAGCCAGAGCCTGAGCATTACCTGCACCGTGAGCGGCTTTAGCCTGACCAACTATGGCGTGCATTGGGTGCGCCAGAGCCCGGGCAAAGGCCTGGAATGGCTGGGCGTGATTTGGAGCGGCGGCAACACCGATTATAACACCCCGTTTACCAGCCGCCTGAGCATTAACAAAGATAACAGCAAAAGCCAGGTGTTTTTTAAAATGAACAGCCTGCAAAGCCAGGATACCGCGATTTATTATTGCGCGCGCGCGCTGACCTATTATGATTATGAATTTGCGTATTGGGGCCAGGGCACCCTGGTGACCGTGAGCGCGGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA] Amino Acid Sequence[spacer (SEQ ID NO: 87)][pLW022 without spacer (SEQ ID NO: 510)](SEQ ID NO: 454)[QGQSGQ][GYRWGCEWNCGGITTGSSGGSGGSGGLSGRSDNHGGGSQTVVTQEPSLTVSPGGTVTLTCRSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGTPARFSGSLLGGKAALTLSGVQPEDEAEYYCALWYSNLWVFGGGTKLTVLGGGGSGGGGSGGGGSEVQLVESGGGLVQPGGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKNTAYLQMNNLKTEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSSGGGGSQVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSQDTAIYYCARALTYYDYEFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]*OPP022: LC C225 3954-1204 Nucleotide Sequence[spacer (SEQ ID NO: 507)][OPP022 without spacer (SEQ ID NO: 509)](SEQ ID NO: 449)[CAAGGCCAGTCTGGCCAG][TGCATCTCACCTCGTGGTTGTCCGGACGGCCCATACGTCATGTACGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGATCCGGTCTGAGCGGCCGTTCCGATAATCATGGCAGTAGCGGTACCCAGATCTTGCTGACCCAGAGCCCGGTGATTCTGAGCGTGAGCCCGGGCGAACGTGTGAGCTTTAGCTGCCGCGCGAGCCAGAGCATTGGCACCAACATTCATTGGTATCAGCAGCGCACCAACGGCAGCCCGCGCCTGCTGATTAAATATGCGAGCGAAAGCATTAGCGGCATTCCGAGCCGCTTTAGCGGCAGCGGCAGCGGCACCGATTTTACCCTGAGCATTAACAGCGTGGAAAGCGAAGATATTGCGGATTATTATTGCCAGCAGAACAACAACTGGCCGACCACCTTTGGCGCGGGCACCAAACTGGAACTGAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG] Amino Acid Sequence[spacer (SEQ ID NO: 87)][OPP022 without spacer (SEQ ID NO: 508)](SEQ ID NO: 450)[QGQSGQ][CISPRGCPDGPYVMYGSSGGSGGSGGSGLSGRSDNHGSSGTQILLTQSPVILSVSPGERVSFSCRASQSIGTNIHWYQQRTNGSPRLLIKYASESISGIPSRFSGSGSGTDFTLSINSVESEDIADYYCQQNNNWPTTFGAGTKLELKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC*]CI011: 3954-1204-C225v5N297Q-15865-CD3LvHv-H-NpLW023: HC C225v5N297Q-15865-CD3LvHv-H-N Nucleotide Sequence[spacer (SEQ ID NO: 507)][pLW023 without spacer (SEQ ID NO: 511)](SEQ ID NO: 451)[CAAGGCCAGTCTGGCCAA][ATGATGTATTGCGGTGGGAATGAGGTGTTGTGCGGGCCGCGGGTTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTCAGACCGTGGTCACACAGGAGCCCTCACTGACAGTGAGCCCTGGCGGGACCGTCACACTGACTTGTCGCAGTTCAACTGGCGCCGTGACTACCAGCAATTACGCTAACTGGGTCCAGCAGAAACCAGGACAGGCACCACGAGGACTGATCGGAGGAACTAATAAGAGAGCACCAGGAACCCCTGCAAGGTTCTCCGGATCTCTGCTGGGGGGAAAAGCCGCTCTGACACTGAGCGGCGTGCAGCCTGAGGACGAAGCTGAGTACTATTGCGCACTGTGGTACTCCAACCTGTGGGTGTTTGGCGGGGGAACTAAGCTGACCGTCCTGGGAGGAGGAGGAAGCGGAGGAGGAGGGAGCGGAGGAGGAGGATCCGAAGTGCAGCTGGTCGAGAGCGGAGGAGGACTGGTGCAGCCAGGAGGATCCCTGAAGCTGTCTTGTGCAGCCAGTGGCTTCACCTTCAACACTTACGCAATGAACTGGGTGCGGCAGGCACCTGGGAAGGGACTGGAATGGGTCGCCCGGATCAGATCTAAATACAATAACTATGCCACCTACTATGCTGACAGTGTGAAGGATAGGTTCACCATTTCACGCGACGATAGCAAAAACACAGCTTATCTGCAGATGAATAACCTGAAGACCGAGGATACAGCAGTGTACTATTGCGTCAGACACGGCAATTTCGGGAACTCTTACGTGAGTTGGTTTGCCTATTGGGGACAGGGGACACTGGTCACCGTCTCCTCAGGAGGTGGTGGATCCCAGGTGCAGCTGAAACAGAGCGGCCCGGGCCTGGTGCAGCCGAGCCAGAGCCTGAGCATTACCTGCACCGTGAGCGGCTTTAGCCTGACCAACTATGGCGTGCATTGGGTGCGCCAGAGCCCGGGCAAAGGCCTGGAATGGCTGGGCGTGATTTGGAGCGGCGGCAACACCGATTATAACACCCCGTTTACCAGCCGCCTGAGCATTAACAAAGATAACAGCAAAAGCCAGGTGTTTTTTAAAATGAACAGCCTGCAAAGCCAGGATACCGCGATTTATTATTGCGCGCGCGCGCTGACCTATTATGATTATGAATTTGCGTATTGGGGCCAGGGCACCCTGGTGACCGTGAGCGCGGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA] Amino Acid Sequence[spacer (SEQ ID NO: 87)][pLW023 without spacer (SEQ ID NO: 512)](SEQ ID NO: 452)[QGQSGQ][MMYCGGNEVLCGPRVGSSGGSGGSGGLSGRSDNHGGGSQTVVTQEPSLTVSPGGTVTLTCRSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGTPARFSGSLLGGKAALTLSGVQPEDEAEYYCALWYSNLWVFGGGTKLTVLGGGGSGGGGSGGGGSEVQLVESGGGLVQPGGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKNTAYLQMNNLKTEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSSGGGGSQVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSQDTAIYYCARALTYYDYEFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]OPP022: LC C225 3954-1204 Nucleotide Sequence[spacer (SEQ ID NO: 507)][OPP022 without spacer (SEQ ID NO: 509)](SEQ ID NO: 449)[CAAGGCCAGTCTGGCCAG][TGCATCTCACCTCGTGGTTGTCCGGACGGCCCATACGTCATGTACGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGATCCGGTCTGAGCGGCCGTTCCGATAATCATGGCAGTAGCGGTACCCAGATCTTGCTGACCCAGAGCCCGGTGATTCTGAGCGTGAGCCCGGGCGAACGTGTGAGCTTTAGCTGCCGCGCGAGCCAGAGCATTGGCACCAACATTCATTGGTATCAGCAGCGCACCAACGGCAGCCCGCGCCTGCTGATTAAATATGCGAGCGAAAGCATTAGCGGCATTCCGAGCCGCTTTAGCGGCAGCGGCAGCGGCACCGATTTTACCCTGAGCATTAACAGCGTGGAAAGCGAAGATATTGCGGATTATTATTGCCAGCAGAACAACAACTGGCCGACCACCTTTGGCGCGGGCACCAAACTGGAACTGAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG] Amino Acid Sequence[spacer (SEQ ID NO: 87)][OPP022 without spacer (SEQ ID NO: 508)](SEQ ID NO: 450)[QGQSGQ][CISPRGCPDGPYVMYGSSGGSGGSGGSGLSGRSDNHGSSGTQILLTQSPVILSVSPGERVSFSCRASQSIGTNIHWYQQRTNGSPRLLIKYASESISGIPSRFSGSGSGTDFTLSINSVESEDIADYYCQQNNNWPTTFGAGTKLELKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC] CI012: C225v5N297QpLW006: C225v5N297Q Nucleotide Sequence (SEQ ID NO: 455)CAGGTGCAGCTGAAACAGAGCGGCCCGGGCCTGGTGCAGCCGAGCCAGAGCCTGAGCATTACCTGCACCGTGAGCGGCTTTAGCCTGACCAACTATGGCGTGCATTGGGTGCGCCAGAGCCCGGGCAAAGGCCTGGAATGGCTGGGCGTGATTTGGAGCGGCGGCAACACCGATTATAACACCCCGTTTACCAGCCGCCTGAGCATTAACAAAGATAACAGCAAAAGCCAGGTGTTTTTTAAAATGAACAGCCTGCAAAGCCAGGATACCGCGATTTATTATTGCGCGCGCGCGCTGACCTATTATGATTATGAATTTGCGTATTGGGGCCAGGGCACCCTGGTGACCGTGAGCGCGGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA  Amino Acid Sequence(SEQ ID NO: 456)QVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSQDTAIYYCARALTYYDYEFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK* OPP021: LC C225 Nucleotide Sequence (SEQ ID NO: 457)GATATCTTGCTGACCCAGAGCCCGGTGATTCTGAGCGTGAGCCCGGGCGAACGTGTGAGCTTTAGCTGCCGCGCGAGCCAGAGCATTGGCACCAACATTCATTGGTATCAGCAGCGCACCAACGGCAGCCCGCGCCTGCTGATTAAATATGCGAGCGAAAGCATTAGCGGCATTCCGAGCCGCTTTAGCGGCAGCGGCAGCGGCACCGATTTTACCCTGAGCATTAACAGCGTGGAAAGCGAAGATATTGCGGATTATTATTGCCAGCAGAACAACAACTGGCCGACCACCTTTGGCGCGGGCACCAAACTGGAAATCAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG  Amino Acid Sequence (SEQ ID NO: 458)DILLTQSPVILSVSPGERVSFSCRASQSIGTNIHWYQQRTNGSPRLLIKYASESISGIPSRFSGSGSGTDFTLSINSVESEDIADYYCQQNNNWPTTFGAGTKLEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC* CI015: C225v5N297Q-CD3LvHv-H-N pLW057: HC C225v5N297Q-CD3LvHv-H-NNucleotide Sequence (SEQ ID NO: 459)CAGACCGTGGTCACACAGGAGCCCTCACTGACAGTGAGCCCTGGCGGGACCGTCACACTGACTTGTCGCAGTTCAACTGGCGCCGTGACTACCAGCAATTACGCTAACTGGGTCCAGCAGAAACCAGGACAGGCACCACGAGGACTGATCGGAGGAACTAATAAGAGAGCACCAGGAACCCCTGCAAGGTTCTCCGGATCTCTGCTGGGGGGAAAAGCCGCTCTGACACTGAGCGGCGTGCAGCCTGAGGACGAAGCTGAGTACTATTGCGCACTGTGGTACTCCAACCTGTGGGTGTTTGGCGGGGGAACTAAGCTGACCGTCCTGGGAGGAGGAGGAAGCGGAGGAGGAGGGAGCGGAGGAGGAGGATCCGAAGTGCAGCTGGTCGAGAGCGGAGGAGGACTGGTGCAGCCAGGAGGATCCCTGAAGCTGTCTTGTGCAGCCAGTGGCTTCACCTTCAACACTTACGCAATGAACTGGGTGCGGCAGGCACCTGGGAAGGGACTGGAATGGGTCGCCCGGATCAGATCTAAATACAATAACTATGCCACCTACTATGCTGACAGTGTGAAGGATAGGTTCACCATTTCACGCGACGATAGCAAAAACACAGCTTATCTGCAGATGAATAACCTGAAGACCGAGGATACAGCAGTGTACTATTGCGTCAGACACGGCAATTTCGGGAACTCTTACGTGAGTTGGTTTGCCTATTGGGGACAGGGGACACTGGTCACCGTCTCCTCAGGAGGTGGTGGATCCCAGGTGCAGCTGAAACAGAGCGGCCCGGGCCTGGTGCAGCCGAGCCAGAGCCTGAGCATTACCTGCACCGTGAGCGGCTTTAGCCTGACCAACTATGGCGTGCATTGGGTGCGCCAGAGCCCGGGCAAAGGCCTGGAATGGCTGGGCGTGATTTGGAGCGGCGGCAACACCGATTATAACACCCCGTTTACCAGCCGCCTGAGCATTAACAAAGATAACAGCAAAAGCCAGGTGTTTTTTAAAATGAACAGCCTGCAAAGCCAGGATACCGCGATTTATTATTGCGCGCGCGCGCTGACCTATTATGATTATGAATTTGCGTATTGGGGCCAGGGCACCCTGGTGACCGTGAGCGCGGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA  Amino Acid Sequence (SEQ ID NO: 460)QTVVTQEPSLTVSPGGTVTLTCRSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGTPARFSGSLLGGKAALTLSGVQPEDEAEYYCALWYSNLWVFGGGTKLTVLGGGGSGGGGSGGGGSEVQLVESGGGLVQPGGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKNTAYLQMNNLKTEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSSGGGGSQVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSQDTAIYYCARALTYYDYEFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK*  OPP021: LC C225 Nucleotide Sequence (SEQ ID NO: 457)GATATCTTGCTGACCCAGAGCCCGGTGATTCTGAGCGTGAGCCCGGGCGAACGTGTGAGCTTTAGCTGCCGCGCGAGCCAGAGCATTGGCACCAACATTCATTGGTATCAGCAGCGCACCAACGGCAGCCCGCGCCTGCTGATTAAATATGCGAGCGAAAGCATTAGCGGCATTCCGAGCCGCTTTAGCGGCAGCGGCAGCGGCACCGATTTTACCCTGAGCATTAACAGCGTGGAAAGCGAAGATATTGCGGATTATTATTGCCAGCAGAACAACAACTGGCCGACCACCTTTGGCGCGGGCACCAAACTGGAAATCAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG  Amino Acid Sequence (SEQ ID NO: 458)DILLTQSPVILSVSPGERVSFSCRASQSIGTNIHWYQQRTNGSPRLLIKYASESISGIPSRFSGSGSGTDFTLSINSVESEDIADYYCQQNNNWPTTFGAGTKLEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC* CI016: 3954-1204-C225v5N297Q-CD3LvHv-H-NpLW057: HC C225v5N297Q-CD3LvHv-H-N Nucleotide Sequence (SEQ ID NO: 459)CAGACCGTGGTCACACAGGAGCCCTCACTGACAGTGAGCCCTGGCGGGACCGTCACACTGACTTGTCGCAGTTCAACTGGCGCCGTGACTACCAGCAATTACGCTAACTGGGTCCAGCAGAAACCAGGACAGGCACCACGAGGACTGATCGGAGGAACTAATAAGAGAGCACCAGGAACCCCTGCAAGGTTCTCCGGATCTCTGCTGGGGGGAAAAGCCGCTCTGACACTGAGCGGCGTGCAGCCTGAGGACGAAGCTGAGTACTATTGCGCACTGTGGTACTCCAACCTGTGGGTGTTTGGCGGGGGAACTAAGCTGACCGTCCTGGGAGGAGGAGGAAGCGGAGGAGGAGGGAGCGGAGGAGGAGGATCCGAAGTGCAGCTGGTCGAGAGCGGAGGAGGACTGGTGCAGCCAGGAGGATCCCTGAAGCTGTCTTGTGCAGCCAGTGGCTTCACCTTCAACACTTACGCAATGAACTGGGTGCGGCAGGCACCTGGGAAGGGACTGGAATGGGTCGCCCGGATCAGATCTAAATACAATAACTATGCCACCTACTATGCTGACAGTGTGAAGGATAGGTTCACCATTTCACGCGACGATAGCAAAAACACAGCTTATCTGCAGATGAATAACCTGAAGACCGAGGATACAGCAGTGTACTATTGCGTCAGACACGGCAATTTCGGGAACTCTTACGTGAGTTGGTTTGCCTATTGGGGACAGGGGACACTGGTCACCGTCTCCTCAGGAGGTGGTGGATCCCAGGTGCAGCTGAAACAGAGCGGCCCGGGCCTGGTGCAGCCGAGCCAGAGCCTGAGCATTACCTGCACCGTGAGCGGCTTTAGCCTGACCAACTATGGCGTGCATTGGGTGCGCCAGAGCCCGGGCAAAGGCCTGGAATGGCTGGGCGTGATTTGGAGCGGCGGCAACACCGATTATAACACCCCGTTTACCAGCCGCCTGAGCATTAACAAAGATAACAGCAAAAGCCAGGTGTTTTTTAAAATGAACAGCCTGCAAAGCCAGGATACCGCGATTTATTATTGCGCGCGCGCGCTGACCTATTATGATTATGAATTTGCGTATTGGGGCCAGGGCACCCTGGTGACCGTGAGCGCGGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA  Amino Acid Sequence (SEQ ID NO: 460)QTVVTQEPSLTVSPGGTVTLTCRSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGTPARFSGSLLGGKAALTLSGVQPEDEAEYYCALWYSNLWVFGGGTKLTVLGGGGSGGGGSGGGGSEVQLVESGGGLVQPGGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKNTAYLQMNNLKTEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSSGGGGSQVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSQDTAIYYCARALTYYDYEFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK* OPP022: LC C225 3954-1204 Nucleotide Sequence[spacer (SEQ ID NO: 507)][OPP022 without spacer (SEQ ID NO: 509)](SEQ ID NO: 449)[CAAGGCCAGTCTGGCCAG][TGCATCTCACCTCGTGGTTGTCCGGACGGCCCATACGTCATGTACGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGATCCGGTCTGAGCGGCCGTTCCGATAATCATGGCAGTAGCGGTACCCAGATCTTGCTGACCCAGAGCCCGGTGATTCTGAGCGTGAGCCCGGGCGAACGTGTGAGCTTTAGCTGCCGCGCGAGCCAGAGCATTGGCACCAACATTCATTGGTATCAGCAGCGCACCAACGGCAGCCCGCGCCTGCTGATTAAATATGCGAGCGAAAGCATTAGCGGCATTCCGAGCCGCTTTAGCGGCAGCGGCAGCGGCACCGATTTTACCCTGAGCATTAACAGCGTGGAAAGCGAAGATATTGCGGATTATTATTGCCAGCAGAACAACAACTGGCCGACCACCTTTGGCGCGGGCACCAAACTGGAACTGAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG] Amino Acid Sequence[spacer (SEQ ID NO: 87)][OPP022 without spacer (SEQ ID NO: 508)](SEQ ID NO: 450)[QGQSGQ][CISPRGCPDGPYVMYGSSGGSGGSGGSGLSGRSDNHGSSGTQILLTQSPVILSVSPGERVSFSCRASQSIGTNIHWYQQRTNGSPRLLIKYASESISGIPSRFSGSGSGTDFTLSINSVESEDIADYYCQQNNNWPTTFGAGTKLELKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC]* CI025: IL6RN297QpLW078: HC IL6RN297Q Nucleotide Sequence (SEQ ID NO: 465)CAGGTGCAGCTGCAGGAGTCCGGACCAGGACTGGTCCGGCCCTCACAGACTCTGAGCCTGACATGCACTGTGTCAGGCTACAGCATCACCTCCGATCACGCCTGGAGCTGGGTCAGGCAGCCACCTGGACGCGGCCTGGAATGGATCGGCTACATTTCTTATAGTGGGATCACCACATACAACCCCTCTCTGAAGAGTCGAGTGACCATTTCCAGAGACAACTCTAAAAATACACTGTATCTGCAGATGAATAGTCTGCGGGCCGAGGATACAGCTGTGTACTATTGTGCACGGTCTCTGGCCAGAACTACCGCTATGGACTATTGGGGGCAGGGAAGCCTGGTGACCGTCAGCTCCGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA  Amino Acid Sequence(SEQ ID NO: 466)QVQLQESGPGLVRPSQTLSLTCTVSGYSITSDHAWSWVRQPPGRGLEWIGYISYSGITTYNPSLKSRVTISRDNSKNTLYLQMNSLRAEDTAVYYCARSLARTTAMDYWGQGSLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK* pLW077: LC IL6R Nucleotide Sequence (SEQ ID NO: 467)GACATCCAGATGACTCAGTCTCCTAGCTCCCTGTCCGCCTCTGTGGGGGACCGAGTCACCATCACATGCAGAGCCAGCCAGGATATTTCTAGTTACCTGAACTGGTATCAGCAGAAGCCCGGAAAAGCACCTAAGCTGCTGATCTACTATACCTCCAGGCTGCACTCTGGCGTGCCCAGTCGGTTCAGTGGCTCAGGGAGCGGAACCGACTTCACTTTTACCATCTCAAGCCTGCAGCCAGAGGATATTGCCACATACTATTGTCAGCAGGGCAATACACTGCCCTACACTTTTGGCCAGGGGACCAAGGTGGAAATCAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG  Amino Acid Sequence (SEQ ID NO: 468)DIQMTQSPSSLSASVGDRVTITCRASQDISSYLNWYQQKPGKAPKLLIYYTSRLHSGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQGNTLPYTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC* CI026: IL6RN297Q-CD3LvHv-H-N pLW083: HC IL6RN297Q-CD3LvHv-H-NNucleotide Sequence (SEQ ID NO: 469)CAGACCGTGGTCACACAGGAGCCCTCACTGACAGTGAGCCCTGGCGGGACCGTCACACTGACTTGTCGCAGTTCAACTGGCGCCGTGACTACCAGCAATTACGCTAACTGGGTCCAGCAGAAACCAGGACAGGCACCACGAGGACTGATCGGAGGAACTAATAAGAGAGCACCAGGAACCCCTGCAAGGTTCTCCGGATCTCTGCTGGGGGGAAAAGCCGCTCTGACACTGAGCGGCGTGCAGCCTGAGGACGAAGCTGAGTACTATTGCGCACTGTGGTACTCCAACCTGTGGGTGTTTGGCGGGGGAACTAAGCTGACCGTCCTGGGAGGAGGAGGAAGCGGAGGAGGAGGGAGCGGAGGAGGAGGATCCGAAGTGCAGCTGGTCGAGAGCGGAGGAGGACTGGTGCAGCCAGGAGGATCCCTGAAGCTGTCTTGTGCAGCCAGTGGCTTCACCTTCAACACTTACGCAATGAACTGGGTGCGGCAGGCACCTGGGAAGGGACTGGAATGGGTCGCCCGGATCAGATCTAAATACAATAACTATGCCACCTACTATGCTGACAGTGTGAAGGATAGGTTCACCATTTCACGCGACGATAGCAAAAACACAGCTTATCTGCAGATGAATAACCTGAAGACCGAGGATACAGCAGTGTACTATTGCGTCAGACACGGCAATTTCGGGAACTCTTACGTGAGTTGGTTTGCCTATTGGGGACAGGGGACACTGGTCACCGTCTCCTCAGGAGGTGGTGGATCCCAGGTGCAGCTGCAGGAGTCCGGACCAGGACTGGTCCGGCCCTCACAGACTCTGAGCCTGACATGCACTGTGTCAGGCTACAGCATCACCTCCGATCACGCCTGGAGCTGGGTCAGGCAGCCACCTGGACGCGGCCTGGAATGGATCGGCTACATTTCTTATAGTGGGATCACCACATACAACCCCTCTCTGAAGAGTCGAGTGACCATTTCCAGAGACAACTCTAAAAATACACTGTATCTGCAGATGAATAGTCTGCGGGCCGAGGATACAGCTGTGTACTATTGTGCACGGTCTCTGGCCAGAACTACCGCTATGGACTATTGGGGGCAGGGAAGCCTGGTGACCGTCAGCTCCGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA  Amino Acid Sequence (SEQ ID NO: 470)QTVVTQEPSLTVSPGGTVTLTCRSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGTPARFSGSLLGGKAALTLSGVQPEDEAEYYCALWYSNLWVFGGGTKLTVLGGGGSGGGGSGGGGSEVQLVESGGGLVQPGGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKNTAYLQMNNLKTEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSSGGGGSQVQLQESGPGLVRPSQTLSLTCTVSGYSITSDHAWSWVRQPPGRGLEWIGYISYSGITTYNPSLKSRVTISRDNSKNTLYLQMNSLRAEDTAVYYCARSLARTTAMDYWGQGSLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK*  pLW077: LC IL6R Nucleotide Sequence (SEQ ID NO: 467)GACATCCAGATGACTCAGTCTCCTAGCTCCCTGTCCGCCTCTGTGGGGGACCGAGTCACCATCACATGCAGAGCCAGCCAGGATATTTCTAGTTACCTGAACTGGTATCAGCAGAAGCCCGGAAAAGCACCTAAGCTGCTGATCTACTATACCTCCAGGCTGCACTCTGGCGTGCCCAGTCGGTTCAGTGGCTCAGGGAGCGGAACCGACTTCACTTTTACCATCTCAAGCCTGCAGCCAGAGGATATTGCCACATACTATTGTCAGCAGGGCAATACACTGCCCTACACTTTTGGCCAGGGGACCAAGGTGGAAATCAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG  Amino Acid Sequence (SEQ ID NO: 468)DIQMTQSPSSLSASVGDRVTITCRASQDISSYLNWYQQKPGKAPKLLIYYTSRLHSGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQGNTLPYTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC* CI027: IL6RN297Q-15865 1204-CD3LvHv-H-NpLW085: HC IL6RN297Q-15865_1204-CD3LvHv-H-N Nucleotide Sequence[spacer (SEQ ID NO: 507)][pLW085 without spacer (SEQ ID NO: 513)](SEQ ID NO: 471)[CAAGGCCAGTCTGGCCAA][ATGATGTATTGCGGTGGGAATGAGGTGTTGTGCGGGCCGCGGGTTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTCAGACCGTGGTCACACAGGAGCCCTCACTGACAGTGAGCCCTGGCGGGACCGTCACACTGACTTGTCGCAGTTCAACTGGCGCCGTGACTACCAGCAATTACGCTAACTGGGTCCAGCAGAAACCAGGACAGGCACCACGAGGACTGATCGGAGGAACTAATAAGAGAGCACCAGGAACCCCTGCAAGGTTCTCCGGATCTCTGCTGGGGGGAAAAGCCGCTCTGACACTGAGCGGCGTGCAGCCTGAGGACGAAGCTGAGTACTATTGCGCACTGTGGTACTCCAACCTGTGGGTGTTTGGCGGGGGAACTAAGCTGACCGTCCTGGGAGGAGGAGGAAGCGGAGGAGGAGGGAGCGGAGGAGGAGGATCCGAAGTGCAGCTGGTCGAGAGCGGAGGAGGACTGGTGCAGCCAGGAGGATCCCTGAAGCTGTCTTGTGCAGCCAGTGGCTTCACCTTCAACACTTACGCAATGAACTGGGTGCGGCAGGCACCTGGGAAGGGACTGGAATGGGTCGCCCGGATCAGATCTAAATACAATAACTATGCCACCTACTATGCTGACAGTGTGAAGGATAGGTTCACCATTTCACGCGACGATAGCAAAAACACAGCTTATCTGCAGATGAATAACCTGAAGACCGAGGATACAGCAGTGTACTATTGCGTCAGACACGGCAATTTCGGGAACTCTTACGTGAGTTGGTTTGCCTATTGGGGACAGGGGACACTGGTCACCGTCTCCTCAGGAGGTGGTGGATCCCAGGTGCAGCTGCAGGAGTCCGGACCAGGACTGGTCCGGCCCTCACAGACTCTGAGCCTGACATGCACTGTGTCAGGCTACAGCATCACCTCCGATCACGCCTGGAGCTGGGTCAGGCAGCCACCTGGACGCGGCCTGGAATGGATCGGCTACATTTCTTATAGTGGGATCACCACATACAACCCCTCTCTGAAGAGTCGAGTGACCATTTCCAGAGACAACTCTAAAAATACACTGTATCTGCAGATGAATAGTCTGCGGGCCGAGGATACAGCTGTGTACTATTGTGCACGGTCTCTGGCCAGAACTACCGCTATGGACTATTGGGGGCAGGGAAGCCTGGTGACCGTCAGCTCCGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA] Amino Acid Sequence[spacer (SEQ ID NO: 87)][pLW085 without spacer (SEQ ID NO: 514)](SEQ ID NO: 472)[QGQSGQ][MMYCGGNEVLCGPRVGSSGGSGGSGGLSGRSDNHGGGSQTVVTQEPSLTVSPGGTVTLTCRSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGTPARFSGSLLGGKAALTLSGVQPEDEAEYYCALWYSNLWVFGGGTKLTVLGGGGSGGGGSGGGGSEVQLVESGGGLVQPGGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKNTAYLQMNNLKTEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSSGGGGSQVQLQESGPGLVRPSQTLSLTCTVSGYSITSDHAWSWVRQPPGRGLEWIGYISYSGITTYNPSLKSRVTISRDNSKNTLYLQMNSLRAEDTAVYYCARSLARTTAMDYWGQGSLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]* pLW077: LC IL6R Nucleotide Sequence (SEQ ID NO: 467)GACATCCAGATGACTCAGTCTCCTAGCTCCCTGTCCGCCTCTGTGGGGGACCGAGTCACCATCACATGCAGAGCCAGCCAGGATATTTCTAGTTACCTGAACTGGTATCAGCAGAAGCCCGGAAAAGCACCTAAGCTGCTGATCTACTATACCTCCAGGCTGCACTCTGGCGTGCCCAGTCGGTTCAGTGGCTCAGGGAGCGGAACCGACTTCACTTTTACCATCTCAAGCCTGCAGCCAGAGGATATTGCCACATACTATTGTCAGCAGGGCAATACACTGCCCTACACTTTTGGCCAGGGGACCAAGGTGGAAATCAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG  Amino Acid Sequence (SEQ ID NO: 468)DIQMTQSPSSLSASVGDRVTITCRASQDISSYLNWYQQKPGKAPKLLIYYTSRLHSGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQGNTLPYTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC* CI029: 4792-1204-1L6RN297Q-15865_1204-CD3LvHv-H-NpLW085: HC IL6RN297Q-15865_1204-CD3LvHv-H-N Nucleotide Sequence[spacer (SEQ ID NO: 507)][pLW085 without spacer (SEQ ID NO: 513)](SEQ ID NO: 471)[CAAGGCCAGTCTGGCCAA][ATGATGTATTGCGGTGGGAATGAGGTGTTGTGCGGGCCGCGGGTTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTCAGACCGTGGTCACACAGGAGCCCTCACTGACAGTGAGCCCTGGCGGGACCGTCACACTGACTTGTCGCAGTTCAACTGGCGCCGTGACTACCAGCAATTACGCTAACTGGGTCCAGCAGAAACCAGGACAGGCACCACGAGGACTGATCGGAGGAACTAATAAGAGAGCACCAGGAACCCCTGCAAGGTTCTCCGGATCTCTGCTGGGGGGAAAAGCCGCTCTGACACTGAGCGGCGTGCAGCCTGAGGACGAAGCTGAGTACTATTGCGCACTGTGGTACTCCAACCTGTGGGTGTTTGGCGGGGGAACTAAGCTGACCGTCCTGGGAGGAGGAGGAAGCGGAGGAGGAGGGAGCGGAGGAGGAGGATCCGAAGTGCAGCTGGTCGAGAGCGGAGGAGGACTGGTGCAGCCAGGAGGATCCCTGAAGCTGTCTTGTGCAGCCAGTGGCTTCACCTTCAACACTTACGCAATGAACTGGGTGCGGCAGGCACCTGGGAAGGGACTGGAATGGGTCGCCCGGATCAGATCTAAATACAATAACTATGCCACCTACTATGCTGACAGTGTGAAGGATAGGTTCACCATTTCACGCGACGATAGCAAAAACACAGCTTATCTGCAGATGAATAACCTGAAGACCGAGGATACAGCAGTGTACTATTGCGTCAGACACGGCAATTTCGGGAACTCTTACGTGAGTTGGTTTGCCTATTGGGGACAGGGGACACTGGTCACCGTCTCCTCAGGAGGTGGTGGATCCCAGGTGCAGCTGCAGGAGTCCGGACCAGGACTGGTCCGGCCCTCACAGACTCTGAGCCTGACATGCACTGTGTCAGGCTACAGCATCACCTCCGATCACGCCTGGAGCTGGGTCAGGCAGCCACCTGGACGCGGCCTGGAATGGATCGGCTACATTTCTTATAGTGGGATCACCACATACAACCCCTCTCTGAAGAGTCGAGTGACCATTTCCAGAGACAACTCTAAAAATACACTGTATCTGCAGATGAATAGTCTGCGGGCCGAGGATACAGCTGTGTACTATTGTGCACGGTCTCTGGCCAGAACTACCGCTATGGACTATTGGGGGCAGGGAAGCCTGGTGACCGTCAGCTCCGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA] Amino Acid Sequence[spacer (SEQ ID NO: 87)][pLW085 without spacer (SEQ ID NO: 514)](SEQ ID NO: 472)[QGQSGQ][MMYCGGNEVLCGPRVGSSGGSGGSGGLSGRSDNHGGGSQTVVTQEPSLTVSPGGTVTLTCRSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGTPARFSGSLLGGKAALTLSGVQPEDEAEYYCALWYSNLWVFGGGTKLTVLGGGGSGGGGSGGGGSEVQLVESGGGLVQPGGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKNTAYLQMNNLKTEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSSGGGGSQVQLQESGPGLVRPSQTLSLTCTVSGYSITSDHAWSWVRQPPGRGLEWIGYISYSGITTYNPSLKSRVTISRDNSKNTLYLQMNSLRAEDTAVYYCARSLARTTAMDYWGQGSLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]*pLW080: LC IL6R 4792 1204 Nucleotide Sequence[spacer (SEQ ID NO: 507)][pLW080 without spacer (SEQ ID NO: 515)](SEQ ID NO: 473)[CAAGGCCAGTCTGGCCAG][TATGGGTCCTGCAGTTGGAACTATGTACACATATTCATGGATTGCGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTGACATCCAGATGACTCAGTCTCCTAGCTCCCTGTCCGCCTCTGTGGGGGACCGAGTCACCATCACATGCAGAGCCAGCCAGGATATTTCTAGTTACCTGAACTGGTATCAGCAGAAGCCCGGAAAAGCACCTAAGCTGCTGATCTACTATACCTCCAGGCTGCACTCTGGCGTGCCCAGTCGGTTCAGTGGCTCAGGGAGCGGAACCGACTTCACTTTTACCATCTCAAGCCTGCAGCCAGAGGATATTGCCACATACTATTGTCAGCAGGGCAATACACTGCCCTACACTTTTGGCCAGGGGACCAAGGTGGAAATCAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG] Amino Acid Sequence[spacer (SEQ ID NO: 87)][pLW080 without spacer (SEQ ID NO: 516)](SEQ ID NO: 474)[QGQSGQ][YGSCSWNYVHIFMDCGSSGGSGGSGGLSGRSDNHGGGSDIQMTQSPSSLSASVGDRVTITCRASQDISSYLNWYQQKPGKAPKLLIYYTSRLHSGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQGNTLPYTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC]*  CI036: 4792-1204-IL6RN297Q-CD3LvHv-H-NpLW083: HC AV1N297Q-CD3LvHv-H-N Nucleotide Sequence (SEQ ID NO: 469)CAGACCGTGGTCACACAGGAGCCCTCACTGACAGTGAGCCCTGGCGGGACCGTCACACTGACTTGTCGCAGTTCAACTGGCGCCGTGACTACCAGCAATTACGCTAACTGGGTCCAGCAGAAACCAGGACAGGCACCACGAGGACTGATCGGAGGAACTAATAAGAGAGCACCAGGAACCCCTGCAAGGTTCTCCGGATCTCTGCTGGGGGGAAAAGCCGCTCTGACACTGAGCGGCGTGCAGCCTGAGGACGAAGCTGAGTACTATTGCGCACTGTGGTACTCCAACCTGTGGGTGTTTGGCGGGGGAACTAAGCTGACCGTCCTGGGAGGAGGAGGAAGCGGAGGAGGAGGGAGCGGAGGAGGAGGATCCGAAGTGCAGCTGGTCGAGAGCGGAGGAGGACTGGTGCAGCCAGGAGGATCCCTGAAGCTGTCTTGTGCAGCCAGTGGCTTCACCTTCAACACTTACGCAATGAACTGGGTGCGGCAGGCACCTGGGAAGGGACTGGAATGGGTCGCCCGGATCAGATCTAAATACAATAACTATGCCACCTACTATGCTGACAGTGTGAAGGATAGGTTCACCATTTCACGCGACGATAGCAAAAACACAGCTTATCTGCAGATGAATAACCTGAAGACCGAGGATACAGCAGTGTACTATTGCGTCAGACACGGCAATTTCGGGAACTCTTACGTGAGTTGGTTTGCCTATTGGGGACAGGGGACACTGGTCACCGTCTCCTCAGGAGGTGGTGGATCCCAGGTGCAGCTGCAGGAGTCCGGACCAGGACTGGTCCGGCCCTCACAGACTCTGAGCCTGACATGCACTGTGTCAGGCTACAGCATCACCTCCGATCACGCCTGGAGCTGGGTCAGGCAGCCACCTGGACGCGGCCTGGAATGGATCGGCTACATTTCTTATAGTGGGATCACCACATACAACCCCTCTCTGAAGAGTCGAGTGACCATTTCCAGAGACAACTCTAAAAATACACTGTATCTGCAGATGAATAGTCTGCGGGCCGAGGATACAGCTGTGTACTATTGTGCACGGTCTCTGGCCAGAACTACCGCTATGGACTATTGGGGGCAGGGAAGCCTGGTGACCGTCAGCTCCGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA  Amino Acid Sequence (SEQ ID NO: 470)QTVVTQEPSLTVSPGGTVTLTCRSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGTPARFSGSLLGGKAALTLSGVQPEDEAEYYCALWYSNLWVFGGGTKLTVLGGGGSGGGGSGGGGSEVQLVESGGGLVQPGGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKNTAYLQMNNLKTEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSSGGGGSQVQLQESGPGLVRPSQTLSLTCTVSGYSITSDHAWSWVRQPPGRGLEWIGYISYSGITTYNPSLKSRVTISRDNSKNTLYLQMNSLRAEDTAVYYCARSLARTTAMDYWGQGSLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK*  pLW080: LC IL6R 4792 1204 Nucleotide Sequence[spacer (SEQ ID NO: 507)][pLW080 without spacer (SEQ ID NO: 515)](SEQ ID NO: 473)[CAAGGCCAGTCTGGCCAG][TATGGGTCCTGCAGTTGGAACTATGTACACATATTCATGGATTGCGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTGACATCCAGATGACTCAGTCTCCTAGCTCCCTGTCCGCCTCTGTGGGGGACCGAGTCACCATCACATGCAGAGCCAGCCAGGATATTTCTAGTTACCTGAACTGGTATCAGCAGAAGCCCGGAAAAGCACCTAAGCTGCTGATCTACTATACCTCCAGGCTGCACTCTGGCGTGCCCAGTCGGTTCAGTGGCTCAGGGAGCGGAACCGACTTCACTTTTACCATCTCAAGCCTGCAGCCAGAGGATATTGCCACATACTATTGTCAGCAGGGCAATACACTGCCCTACACTTTTGGCCAGGGGACCAAGGTGGAAATCAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG] Amino Acid Sequence[spacer (SEQ ID NO: 87)][pLW080 without spacer (SEQ ID NO: 516)](SEQ ID NO: 474)[QGQSGQ][YGSCSWNYVHIFMDCGSSGGSGGSGGLSGRSDNHGGGSDIQMTQSPSSLSASVGDRVTITCRASQDISSYLNWYQQKPGKAPKLLIYYTSRLHSGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQGNTLPYTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC]* CI039: 3954-2001-C225v5 N297Q-15865-2001-CD3LvHv-H-NpLW101: HC C225v5N297Q-15865-2001-CD3LvHv-H-N Nucleotide Sequence[spacer (SEQ ID NO: 507)][pLW101 without spacer (SEQ ID NO: 517)](SEQ ID NO: 479)[CAAGGCCAGTCTGGCCAA][ATGATGTATTGCGGTGGGAATGAGGTGTTGTGCGGGCCGCGGGTTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTATCTCTTCCGGACTGCTGTCCGGCAGATCCGACAATCACGGCGGCGGTTCTCAGACCGTGGTCACACAGGAGCCCTCACTGACAGTGAGCCCTGGCGGGACCGTCACACTGACTTGTCGCAGTTCAACTGGCGCCGTGACTACCAGCAATTACGCTAACTGGGTCCAGCAGAAACCAGGACAGGCACCACGAGGACTGATCGGAGGAACTAATAAGAGAGCACCAGGAACCCCTGCAAGGTTCTCCGGATCTCTGCTGGGGGGAAAAGCCGCTCTGACACTGAGCGGCGTGCAGCCTGAGGACGAAGCTGAGTACTATTGCGCACTGTGGTACTCCAACCTGTGGGTGTTTGGCGGGGGAACTAAGCTGACCGTCCTGGGAGGAGGAGGAAGCGGAGGAGGAGGGAGCGGAGGAGGAGGATCCGAAGTGCAGCTGGTCGAGAGCGGAGGAGGACTGGTGCAGCCAGGAGGATCCCTGAAGCTGTCTTGTGCAGCCAGTGGCTTCACCTTCAACACTTACGCAATGAACTGGGTGCGGCAGGCACCTGGGAAGGGACTGGAATGGGTCGCCCGGATCAGATCTAAATACAATAACTATGCCACCTACTATGCTGACAGTGTGAAGGATAGGTTCACCATTTCACGCGACGATAGCAAAAACACAGCTTATCTGCAGATGAATAACCTGAAGACCGAGGATACAGCAGTGTACTATTGCGTCAGACACGGCAATTTCGGGAACTCTTACGTGAGTTGGTTTGCCTATTGGGGACAGGGGACACTGGTCACCGTCTCCTCAGGAGGTGGTGGATCCCAGGTGCAGCTGAAACAGAGCGGCCCGGGCCTGGTGCAGCCGAGCCAGAGCCTGAGCATTACCTGCACCGTGAGCGGCTTTAGCCTGACCAACTATGGCGTGCATTGGGTGCGCCAGAGCCCGGGCAAAGGCCTGGAATGGCTGGGCGTGATTTGGAGCGGCGGCAACACCGATTATAACACCCCGTTTACCAGCCGCCTGAGCATTAACAAAGATAACAGCAAAAGCCAGGTGTTTTTTAAAATGAACAGCCTGCAAAGCCAGGATACCGCGATTTATTATTGCGCGCGCGCGCTGACCTATTATGATTATGAATTTGCGTATTGGGGCCAGGGCACCCTGGTGACCGTGAGCGCGGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA] Amino Acid Sequence[spacer (SEQ ID NO: 87)][pLW101 without spacer (SEQ ID NO: 518)](SEQ ID NO: 480)[QGQSGQ][MMYCGGNEVLCGPRVGSSGGSGGSGGISSGLLSGRSDNHGGGSQTVVTQEPSLTVSPGGTVTLTCRSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGTPARFSGSLLGGKAALTLSGVQPEDEAEYYCALWYSNLWVFGGGTKLTVLGGGGSGGGGSGGGGSEVQLVESGGGLVQPGGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKNTAYLQMNNLKTEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSSGGGGSQVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSQDTAIYYCARALTYYDYEFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]*LC C225-3954-2001 Nucleotide Sequence[spacer (SEQ ID NO: 507)][3954-2001-C225v5 without spacer (SEQ ID NO: 519)](SEQ ID NO: 481)[CAAGGCCAGTCTGGCCAG][TGCATCTCACCTCGTGGTTGTCCGGACGGCCCATACGTCATGTACGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGATCCGGTATTAGCAGTGGTCTGTTAAGCGGTCGTAGCGATAATCATGGCAGTAGCGGTACCCAGATCTTGCTGACCCAGAGCCCGGTGATTCTGAGCGTGAGCCCGGGCGAACGTGTGAGCTTTAGCTGCCGCGCGAGCCAGAGCATTGGCACCAACATTCATTGGTATCAGCAGCGCACCAACGGCAGCCCGCGCCTGCTGATTAAATATGCGAGCGAAAGCATTAGCGGCATTCCGAGCCGCTTTAGCGGCAGCGGCAGCGGCACCGATTTTACCCTGAGCATTAACAGCGTGGAAAGCGAAGATATTGCGGATTATTATTGCCAGCAGAACAACAACTGGCCGACCACCTTTGGCGCGGGCACCAAACTGGAACTGAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG] Amino Acid Sequence[spacer (SEQ ID NO: 87)][pLW080 without spacer (SEQ ID NO: 520)](SEQ ID NO: 482)[QGQSGQ][CISPRGCPDGPYVMYGSSGGSGGSGGSGISSGLLSGRSDNHGSSGTQILLTQSPVILSVSPGERVSFSCRASQSIGTNIHWYQQRTNGSPRLLIKYASESISGIPSRFSGSGSGTDFTLSINSVESEDIADYYCQQNNNWPTTFGAGTKLELKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC]* CI040: 3954-2001-C225v5 N297Q-CD3LvHv-H-NpLW057: HC C225v5N297Q-CD3LvHv-H-N Nucleotide Sequence (SEQ ID NO: 479)CAGACCGTGGTCACACAGGAGCCCTCACTGACAGTGAGCCCTGGCGGGACCGTCACACTGACTTGTCGCAGTTCAACTGGCGCCGTGACTACCAGCAATTACGCTAACTGGGTCCAGCAGAAACCAGGACAGGCACCACGAGGACTGATCGGAGGAACTAATAAGAGAGCACCAGGAACCCCTGCAAGGTTCTCCGGATCTCTGCTGGGGGGAAAAGCCGCTCTGACACTGAGCGGCGTGCAGCCTGAGGACGAAGCTGAGTACTATTGCGCACTGTGGTACTCCAACCTGTGGGTGTTTGGCGGGGGAACTAAGCTGACCGTCCTGGGAGGAGGAGGAAGCGGAGGAGGAGGGAGCGGAGGAGGAGGATCCGAAGTGCAGCTGGTCGAGAGCGGAGGAGGACTGGTGCAGCCAGGAGGATCCCTGAAGCTGTCTTGTGCAGCCAGTGGCTTCACCTTCAACACTTACGCAATGAACTGGGTGCGGCAGGCACCTGGGAAGGGACTGGAATGGGTCGCCCGGATCAGATCTAAATACAATAACTATGCCACCTACTATGCTGACAGTGTGAAGGATAGGTTCACCATTTCACGCGACGATAGCAAAAACACAGCTTATCTGCAGATGAATAACCTGAAGACCGAGGATACAGCAGTGTACTATTGCGTCAGACACGGCAATTTCGGGAACTCTTACGTGAGTTGGTTTGCCTATTGGGGACAGGGGACACTGGTCACCGTCTCCTCAGGAGGTGGTGGATCCCAGGTGCAGCTGAAACAGAGCGGCCCGGGCCTGGTGCAGCCGAGCCAGAGCCTGAGCATTACCTGCACCGTGAGCGGCTTTAGCCTGACCAACTATGGCGTGCATTGGGTGCGCCAGAGCCCGGGCAAAGGCCTGGAATGGCTGGGCGTGATTTGGAGCGGCGGCAACACCGATTATAACACCCCGTTTACCAGCCGCCTGAGCATTAACAAAGATAACAGCAAAAGCCAGGTGTTTTTTAAAATGAACAGCCTGCAAAGCCAGGATACCGCGATTTATTATTGCGCGCGCGCGCTGACCTATTATGATTATGAATTTGCGTATTGGGGCCAGGGCACCCTGGTGACCGTGAGCGCGGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA  Amino Acid Sequence (SEQ ID NO: 480)QTVVTQEPSLTVSPGGTVTLTCRSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGTPARFSGSLLGGKAALTLSGVQPEDEAEYYCALWYSNLWVFGGGTKLTVLGGGGSGGGGSGGGGSEVQLVESGGGLVQPGGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKNTAYLQMNNLKTEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSSGGGGSQVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSQDTAIYYCARALTYYDYEFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK*  LC C225-3954-2001 Nucleotide Sequence[spacer (SEQ ID NO: 507)][3954-2001-C225v5without spacer (SEQ ID NO: 519)](SEQ ID NO: 481)[CAAGGCCAGTCTGGCCAG][TGCATCTCACCTCGTGGTTGTCCGGACGGCCCATACGTCATGTACGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGATCCGGTATTAGCAGTGGTCTGTTAAGCGGTCGTAGCGATAATCATGGCAGTAGCGGTACCCAGATCTTGCTGACCCAGAGCCCGGTGATTCTGAGCGTGAGCCCGGGCGAACGTGTGAGCTTTAGCTGCCGCGCGAGCCAGAGCATTGGCACCAACATTCATTGGTATCAGCAGCGCACCAACGGCAGCCCGCGCCTGCTGATTAAATATGCGAGCGAAAGCATTAGCGGCATTCCGAGCCGCTTTAGCGGCAGCGGCAGCGGCACCGATTTTACCCTGAGCATTAACAGCGTGGAAAGCGAAGATATTGCGGATTATTATTGCCAGCAGAACAACAACTGGCCGACCACCTTTGGCGCGGGCACCAAACTGGAACTGAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG] Amino Acid Sequence[spacer (SEQ ID NO: 87)][pLW080 without spacer (SEQ ID NO: 520)](SEQ ID NO: 482)[QGQSGQ][CISPRGCPDGPYVMYGSSGGSGGSGGSGISSGLLSGRSDNHGSSGTQILLTQSPVILSVSPGERVSFSCRASQSIGTNIHWYQQRTNGSPRLLIKYASESISGIPSRFSGSGSGTDFTLSINSVESEDIADYYCQQNNNWPTTFGAGTKLELKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC]*CI021: C225v5N297Q-15003-CD3HvLv-H-NpLW047: HC C225v5N297Q-15003-CD3HvLv-H-N Nucleotide Sequence[spacer (SEQ ID NO: 507)][pLW047 without spacer (SEQ ID NO: 522)](SEQ ID NO: 461)[CAAGGCCAGTCTGGCCAA][GGTTATCGGTGGGGTTGCGAGTGGAATTGCGGTGGGATTACTACTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTGAGGTGCAGCTGGTCGAGTCTGGAGGAGGATTGGTGCAGCCTGGAGGGTCATTGAAACTCTCATGTGCAGCCTCTGGATTCACCTTCAATAAGTACGCCATGAACTGGGTCCGCCAGGCTCCAGGAAAGGGTTTGGAATGGGTTGCTCGCATAAGAAGTAAATATAATAATTATGCAACATATTATGCCGATTCAGTGAAAGACAGGTTCACCATCTCCAGAGATGATTCAAAAAACACTGCCTATCTACAAATGAACAACTTGAAAACTGAGGACACTGCCGTGTACTACTGTGTGAGACATGGGAACTTCGGTAATAGCTACATATCCTACTGGGCTTACTGGGGCCAAGGGACTCTGGTCACCGTCTCCTCAGGTGGTGGTGGTTCTGGCGGCGGCGGCTCCGGTGGTGGTGGTTCTCAGACTGTTGTGACTCAGGAACCTTCACTCACCGTATCACCTGGTGGAACAGTCACACTCACTTGTGGCTCCTCGACTGGGGCTGTTACATCTGGCTACTACCCAAACTGGGTCCAACAAAAACCAGGTCAGGCACCCCGTGGTCTAATAGGTGGGACTAAGTTCCTCGCCCCCGGTACTCCTGCCAGATTCTCAGGCTCCCTGCTTGGAGGCAAGGCTGCCCTCACCCTCTCAGGGGTACAGCCAGAGGATGAGGCAGAATATTACTGTGCTCTATGGTACAGCAACCGCTGGGTGTTCGGTGGAGGAACCAAACTGACTGTCCTAGGAGGTGGTGGATCCCAGGTGCAGCTGAAACAGAGCGGCCCGGGCCTGGTGCAGCCGAGCCAGAGCCTGAGCATTACCTGCACCGTGAGCGGCTTTAGCCTGACCAACTATGGCGTGCATTGGGTGCGCCAGAGCCCGGGCAAAGGCCTGGAATGGCTGGGCGTGATTTGGAGCGGCGGCAACACCGATTATAACACCCCGTTTACCAGCCGCCTGAGCATTAACAAAGATAACAGCAAAAGCCAGGTGTTTTTTAAAATGAACAGCCTGCAAAGCCAGGATACCGCGATTTATTATTGCGCGCGCGCGCTGACCTATTATGATTATGAATTTGCGTATTGGGGCCAGGGCACCCTGGTGACCGTGAGCGCGGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA] Amino Acid Sequence[spacer (SEQ ID NO: 87)][pLW047 without spacer (SEQ ID NO: 524)](SEQ ID NO: 462)[QGQSGQ][GYRWGCEWNCGGITTGSSGGSGGSGGLSGRSDNHGGGSEVQLVESGGGLVQPGGSLKLSCAASGFTFNKYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKNTAYLQMNNLKTEDTAVYYCVRHGNFGNSYISYWAYWGQGTLVTVSSGGGGSGGGGSGGGGSQTVVTQEPSLTVSPGGTVTLTCGSSTGAVTSGYYPNWVQQKPGQAPRGLIGGTKFLAPGTPARFSGSLLGGKAALTLSGVQPEDEAEYYCALWYSNRWVFGGGTKLTVLGGGGSQVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSQDTAIYYCARALTYYDYEFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]* OPP021: LC C225 Nucleotide Sequence (SEQ ID NO: 457)GATATCTTGCTGACCCAGAGCCCGGTGATTCTGAGCGTGAGCCCGGGCGAACGTGTGAGCTTTAGCTGCCGCGCGAGCCAGAGCATTGGCACCAACATTCATTGGTATCAGCAGCGCACCAACGGCAGCCCGCGCCTGCTGATTAAATATGCGAGCGAAAGCATTAGCGGCATTCCGAGCCGCTTTAGCGGCAGCGGCAGCGGCACCGATTTTACCCTGAGCATTAACAGCGTGGAAAGCGAAGATATTGCGGATTATTATTGCCAGCAGAACAACAACTGGCCGACCACCTTTGGCGCGGGCACCAAACTGGAAATCAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG  Amino Acid Sequence (SEQ ID NO: 458)DILLTQSPVILSVSPGERVSFSCRASQSIGTNIHWYQQRTNGSPRLLIKYASESISGIPSRFSGSGSGTDFTLSINSVESEDIADYYCQQNNNWPTTFGAGTKLEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC* CI022: C225v5N297Q-15865-CD3HvLv-H-NpLW048: HC C225v5N297Q-15865-CD3HvLv-H-N Nucleotide Sequence[spacer (SEQ ID NO: 507)][pLW048 without spacer (SEQ ID NO: 525)](SEQ ID NO: 463)[CAAGGCCAGTCTGGCCAA][ATGATGTATTGCGGTGGGAATGAGGTGTTGTGCGGGCCGCGGGTTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTGAGGTGCAGCTGGTCGAGTCTGGAGGAGGATTGGTGCAGCCTGGAGGGTCATTGAAACTCTCATGTGCAGCCTCTGGATTCACCTTCAATAAGTACGCCATGAACTGGGTCCGCCAGGCTCCAGGAAAGGGTTTGGAATGGGTTGCTCGCATAAGAAGTAAATATAATAATTATGCAACATATTATGCCGATTCAGTGAAAGACAGGTTCACCATCTCCAGAGATGATTCAAAAAACACTGCCTATCTACAAATGAACAACTTGAAAACTGAGGACACTGCCGTGTACTACTGTGTGAGACATGGGAACTTCGGTAATAGCTACATATCCTACTGGGCTTACTGGGGCCAAGGGACTCTGGTCACCGTCTCCTCAGGTGGTGGTGGTTCTGGCGGCGGCGGCTCCGGTGGTGGTGGTTCTCAGACTGTTGTGACTCAGGAACCTTCACTCACCGTATCACCTGGTGGAACAGTCACACTCACTTGTGGCTCCTCGACTGGGGCTGTTACATCTGGCTACTACCCAAACTGGGTCCAACAAAAACCAGGTCAGGCACCCCGTGGTCTAATAGGTGGGACTAAGTTCCTCGCCCCCGGTACTCCTGCCAGATTCTCAGGCTCCCTGCTTGGAGGCAAGGCTGCCCTCACCCTCTCAGGGGTACAGCCAGAGGATGAGGCAGAATATTACTGTGCTCTATGGTACAGCAACCGCTGGGTGTTCGGTGGAGGAACCAAACTGACTGTCCTAGGAGGTGGTGGATCCCAGGTGCAGCTGAAACAGAGCGGCCCGGGCCTGGTGCAGCCGAGCCAGAGCCTGAGCATTACCTGCACCGTGAGCGGCTTTAGCCTGACCAACTATGGCGTGCATTGGGTGCGCCAGAGCCCGGGCAAAGGCCTGGAATGGCTGGGCGTGATTTGGAGCGGCGGCAACACCGATTATAACACCCCGTTTACCAGCCGCCTGAGCATTAACAAAGATAACAGCAAAAGCCAGGTGTTTTTTAAAATGAACAGCCTGCAAAGCCAGGATACCGCGATTTATTATTGCGCGCGCGCGCTGACCTATTATGATTATGAATTTGCGTATTGGGGCCAGGGCACCCTGGTGACCGTGAGCGCGGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA] Amino Acid Sequence[spacer (SEQ ID NO: 87)][pLW048 without spacer (SEQ ID NO: 526)](SEQ ID NO: 464)[QGQSGQ][MMYCGGNEVLCGPRVGSSGGSGGSGGLSGRSDNHGGGSEVQLVESGGGLVQPGGSLKLSCAASGFTFNKYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKNTAYLQMNNLKTEDTAVYYCVRHGNFGNSYISYWAYWGQGTLVTVSSGGGGSGGGGSGGGGSQTVVTQEPSLTVSPGGTVTLTCGSSTGAVTSGYYPNWVQQKPGQAPRGLIGGTKFLAPGTPARFSGSLLGGKAALTLSGVQPEDEAEYYCALWYSNRWVFGGGTKLTVLGGGGSQVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSQDTAIYYCARALTYYDYEFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]* OPP021: LC C225 Nucleotide Sequence (SEQ ID NO: 457)GATATCTTGCTGACCCAGAGCCCGGTGATTCTGAGCGTGAGCCCGGGCGAACGTGTGAGCTTTAGCTGCCGCGCGAGCCAGAGCATTGGCACCAACATTCATTGGTATCAGCAGCGCACCAACGGCAGCCCGCGCCTGCTGATTAAATATGCGAGCGAAAGCATTAGCGGCATTCCGAGCCGCTTTAGCGGCAGCGGCAGCGGCACCGATTTTACCCTGAGCATTAACAGCGTGGAAAGCGAAGATATTGCGGATTATTATTGCCAGCAGAACAACAACTGGCCGACCACCTTTGGCGCGGGCACCAAACTGGAAATCAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG  Amino Acid Sequence (SEQ ID NO: 458)DILLTQSPVILSVSPGERVSFSCRASQSIGTNIHWYQQRTNGSPRLLIKYASESISGIPSRFSGSGSGTDFTLSINSVESEDIADYYCQQNNNWPTTFGAGTKLEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC* CI023: 3954-1204-C225v5N297Q-15003-CD3HvLv-H-NpLW047: HC C225v5N297Q-15003-CD3HvLv-H-N Nucleotide Sequence[spacer (SEQ ID NO: 507)][pLW047 without spacer (SEQ ID NO: 522)](SEQ ID NO: 461)[CAAGGCCAGTCTGGCCAA][GGTTATCGGTGGGGTTGCGAGTGGAATTGCGGTGGGATTACTACTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTGAGGTGCAGCTGGTCGAGTCTGGAGGAGGATTGGTGCAGCCTGGAGGGTCATTGAAACTCTCATGTGCAGCCTCTGGATTCACCTTCAATAAGTACGCCATGAACTGGGTCCGCCAGGCTCCAGGAAAGGGTTTGGAATGGGTTGCTCGCATAAGAAGTAAATATAATAATTATGCAACATATTATGCCGATTCAGTGAAAGACAGGTTCACCATCTCCAGAGATGATTCAAAAAACACTGCCTATCTACAAATGAACAACTTGAAAACTGAGGACACTGCCGTGTACTACTGTGTGAGACATGGGAACTTCGGTAATAGCTACATATCCTACTGGGCTTACTGGGGCCAAGGGACTCTGGTCACCGTCTCCTCAGGTGGTGGTGGTTCTGGCGGCGGCGGCTCCGGTGGTGGTGGTTCTCAGACTGTTGTGACTCAGGAACCTTCACTCACCGTATCACCTGGTGGAACAGTCACACTCACTTGTGGCTCCTCGACTGGGGCTGTTACATCTGGCTACTACCCAAACTGGGTCCAACAAAAACCAGGTCAGGCACCCCGTGGTCTAATAGGTGGGACTAAGTTCCTCGCCCCCGGTACTCCTGCCAGATTCTCAGGCTCCCTGCTTGGAGGCAAGGCTGCCCTCACCCTCTCAGGGGTACAGCCAGAGGATGAGGCAGAATATTACTGTGCTCTATGGTACAGCAACCGCTGGGTGTTCGGTGGAGGAACCAAACTGACTGTCCTAGGAGGTGGTGGATCCCAGGTGCAGCTGAAACAGAGCGGCCCGGGCCTGGTGCAGCCGAGCCAGAGCCTGAGCATTACCTGCACCGTGAGCGGCTTTAGCCTGACCAACTATGGCGTGCATTGGGTGCGCCAGAGCCCGGGCAAAGGCCTGGAATGGCTGGGCGTGATTTGGAGCGGCGGCAACACCGATTATAACACCCCGTTTACCAGCCGCCTGAGCATTAACAAAGATAACAGCAAAAGCCAGGTGTTTTTTAAAATGAACAGCCTGCAAAGCCAGGATACCGCGATTTATTATTGCGCGCGCGCGCTGACCTATTATGATTATGAATTTGCGTATTGGGGCCAGGGCACCCTGGTGACCGTGAGCGCGGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA] Amino Acid Sequence[spacer (SEQ ID NO: 87)][pLW047 without spacer (SEQ ID NO: 524)](SEQ ID NO: 462)[QGQSGQ][GYRWGCEWNCGGITTGSSGGSGGSGGLSGRSDNHGGGSEVQLVESGGGLVQPGGSLKLSCAASGFTFNKYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKNTAYLQMNNLKTEDTAVYYCVRHGNFGNSYISYWAYWGQGTLVTVSSGGGGSGGGGSGGGGSQTVVTQEPSLTVSPGGTVTLTCGSSTGAVTSGYYPNWVQQKPGQAPRGLIGGTKFLAPGTPARFSGSLLGGKAALTLSGVQPEDEAEYYCALWYSNRWVFGGGTKLTVLGGGGSQVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSQDTAIYYCARALTYYDYEFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]*OPP022: LC C225 3954-1204 Nucleotide Sequence[spacer (SEQ ID NO: 507)][OPP022 without spacer (SEQ ID NO: 509)](SEQ ID NO: 449)[CAAGGCCAGTCTGGCCAG][TGCATCTCACCTCGTGGTTGTCCGGACGGCCCATACGTCATGTACGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGATCCGGTCTGAGCGGCCGTTCCGATAATCATGGCAGTAGCGGTACCCAGATCTTGCTGACCCAGAGCCCGGTGATTCTGAGCGTGAGCCCGGGCGAACGTGTGAGCTTTAGCTGCCGCGCGAGCCAGAGCATTGGCACCAACATTCATTGGTATCAGCAGCGCACCAACGGCAGCCCGCGCCTGCTGATTAAATATGCGAGCGAAAGCATTAGCGGCATTCCGAGCCGCTTTAGCGGCAGCGGCAGCGGCACCGATTTTACCCTGAGCATTAACAGCGTGGAAAGCGAAGATATTGCGGATTATTATTGCCAGCAGAACAACAACTGGCCGACCACCTTTGGCGCGGGCACCAAACTGGAACTGAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG] Amino Acid Sequence[spacer (SEQ ID NO: 87)][OPP022 without spacer (SEQ ID NO: 508)](SEQ ID NO: 450)[QGQSGQ][CISPRGCPDGPYVMYGSSGGSGGSGGSGLSGRSDNHGSSGTQILLTQSPVILSVSPGERVSFSCRASQSIGTNIHWYQQRTNGSPRLLIKYASESISGIPSRFSGSGSGTDFTLSINSVESEDIADYYCQQNNNWPTTFGAGTKLELKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC]*CI024: 3954-1204-C225v5N297Q-15865-CD3HvLv-H-NpLW048: HC C225v5N297Q-15865-CD3HvLv-H-N Nucleotide Sequence[spacer (SEQ ID NO: 507)][pLW048 without spacer (SEQ ID NO: 525)](SEQ ID NO: 463)[CAAGGCCAGTCTGGCCAA][ATGATGTATTGCGGTGGGAATGAGGTGTTGTGCGGGCCGCGGGTTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTGAGGTGCAGCTGGTCGAGTCTGGAGGAGGATTGGTGCAGCCTGGAGGGTCATTGAAACTCTCATGTGCAGCCTCTGGATTCACCTTCAATAAGTACGCCATGAACTGGGTCCGCCAGGCTCCAGGAAAGGGTTTGGAATGGGTTGCTCGCATAAGAAGTAAATATAATAATTATGCAACATATTATGCCGATTCAGTGAAAGACAGGTTCACCATCTCCAGAGATGATTCAAAAAACACTGCCTATCTACAAATGAACAACTTGAAAACTGAGGACACTGCCGTGTACTACTGTGTGAGACATGGGAACTTCGGTAATAGCTACATATCCTACTGGGCTTACTGGGGCCAAGGGACTCTGGTCACCGTCTCCTCAGGTGGTGGTGGTTCTGGCGGCGGCGGCTCCGGTGGTGGTGGTTCTCAGACTGTTGTGACTCAGGAACCTTCACTCACCGTATCACCTGGTGGAACAGTCACACTCACTTGTGGCTCCTCGACTGGGGCTGTTACATCTGGCTACTACCCAAACTGGGTCCAACAAAAACCAGGTCAGGCACCCCGTGGTCTAATAGGTGGGACTAAGTTCCTCGCCCCCGGTACTCCTGCCAGATTCTCAGGCTCCCTGCTTGGAGGCAAGGCTGCCCTCACCCTCTCAGGGGTACAGCCAGAGGATGAGGCAGAATATTACTGTGCTCTATGGTACAGCAACCGCTGGGTGTTCGGTGGAGGAACCAAACTGACTGTCCTAGGAGGTGGTGGATCCCAGGTGCAGCTGAAACAGAGCGGCCCGGGCCTGGTGCAGCCGAGCCAGAGCCTGAGCATTACCTGCACCGTGAGCGGCTTTAGCCTGACCAACTATGGCGTGCATTGGGTGCGCCAGAGCCCGGGCAAAGGCCTGGAATGGCTGGGCGTGATTTGGAGCGGCGGCAACACCGATTATAACACCCCGTTTACCAGCCGCCTGAGCATTAACAAAGATAACAGCAAAAGCCAGGTGTTTTTTAAAATGAACAGCCTGCAAAGCCAGGATACCGCGATTTATTATTGCGCGCGCGCGCTGACCTATTATGATTATGAATTTGCGTATTGGGGCCAGGGCACCCTGGTGACCGTGAGCGCGGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA] Amino Acid Sequence[spacer (SEQ ID NO: 87)][pLW048 without spacer (SEQ ID NO: 526)](SEQ ID NO: 464)[QGQSGQ][MMYCGGNEVLCGPRVGSSGGSGGSGGLSGRSDNHGGGSEVQLVESGGGLVQPGGSLKLSCAASGFTFNKYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKNTAYLQMNNLKTEDTAVYYCVRHGNFGNSYISYWAYWGQGTLVTVSSGGGGSGGGGSGGGGSQTVVTQEPSLTVSPGGTVTLTCGSSTGAVTSGYYPNWVQQKPGQAPRGLIGGTKFLAPGTPARFSGSLLGGKAALTLSGVQPEDEAEYYCALWYSNRWVFGGGTKLTVLGGGGSQVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSQDTAIYYCARALTYYDYEFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]*OPP022: LC C225 3954-1204 Nucleotide Sequence[spacer (SEQ ID NO: 507)][OPP022 without spacer (SEQ ID NO: 509)](SEQ ID NO: 449)[CAAGGCCAGTCTGGCCAG][TGCATCTCACCTCGTGGTTGTCCGGACGGCCCATACGTCATGTACGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGATCCGGTCTGAGCGGCCGTTCCGATAATCATGGCAGTAGCGGTACCCAGATCTTGCTGACCCAGAGCCCGGTGATTCTGAGCGTGAGCCCGGGCGAACGTGTGAGCTTTAGCTGCCGCGCGAGCCAGAGCATTGGCACCAACATTCATTGGTATCAGCAGCGCACCAACGGCAGCCCGCGCCTGCTGATTAAATATGCGAGCGAAAGCATTAGCGGCATTCCGAGCCGCTTTAGCGGCAGCGGCAGCGGCACCGATTTTACCCTGAGCATTAACAGCGTGGAAAGCGAAGATATTGCGGATTATTATTGCCAGCAGAACAACAACTGGCCGACCACCTTTGGCGCGGGCACCAAACTGGAACTGAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG] Amino Acid Sequence[spacer (SEQ ID NO: 87)][OPP022 without spacer (SEQ ID NO: 508)](SEQ ID NO: 450)[QGQSGQ][CISPRGCPDGPYVMYGSSGGSGGSGGSGLSGRSDNHGSSGTQILLTQSPVILSVSPGERVSFSCRASQSIGTNIHWYQQRTNGSPRLLIKYASESISGIPSRFSGSGSGTDFTLSINSVESEDIADYYCQQNNNWPTTFGAGTKLELKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC]*CI028: IL6R 4792 Nsub N297Q-15865 1204-CD3LvHv-H-NpLW085: HC IL6RN297Q-15865 1204-CD3LvHv-H-N Nucleotide Sequence[spacer (SEQ ID NO: 507)][pLW085 without spacer (SEQ ID NO: 513)](SEQ ID NO: 471)[CAAGGCCAGTCTGGCCAA][ATGATGTATTGCGGTGGGAATGAGGTGTTGTGCGGGCCGCGGGTTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTCAGACCGTGGTCACACAGGAGCCCTCACTGACAGTGAGCCCTGGCGGGACCGTCACACTGACTTGTCGCAGTTCAACTGGCGCCGTGACTACCAGCAATTACGCTAACTGGGTCCAGCAGAAACCAGGACAGGCACCACGAGGACTGATCGGAGGAACTAATAAGAGAGCACCAGGAACCCCTGCAAGGTTCTCCGGATCTCTGCTGGGGGGAAAAGCCGCTCTGACACTGAGCGGCGTGCAGCCTGAGGACGAAGCTGAGTACTATTGCGCACTGTGGTACTCCAACCTGTGGGTGTTTGGCGGGGGAACTAAGCTGACCGTCCTGGGAGGAGGAGGAAGCGGAGGAGGAGGGAGCGGAGGAGGAGGATCCGAAGTGCAGCTGGTCGAGAGCGGAGGAGGACTGGTGCAGCCAGGAGGATCCCTGAAGCTGTCTTGTGCAGCCAGTGGCTTCACCTTCAACACTTACGCAATGAACTGGGTGCGGCAGGCACCTGGGAAGGGACTGGAATGGGTCGCCCGGATCAGATCTAAATACAATAACTATGCCACCTACTATGCTGACAGTGTGAAGGATAGGTTCACCATTTCACGCGACGATAGCAAAAACACAGCTTATCTGCAGATGAATAACCTGAAGACCGAGGATACAGCAGTGTACTATTGCGTCAGACACGGCAATTTCGGGAACTCTTACGTGAGTTGGTTTGCCTATTGGGGACAGGGGACACTGGTCACCGTCTCCTCAGGAGGTGGTGGATCCCAGGTGCAGCTGCAGGAGTCCGGACCAGGACTGGTCCGGCCCTCACAGACTCTGAGCCTGACATGCACTGTGTCAGGCTACAGCATCACCTCCGATCACGCCTGGAGCTGGGTCAGGCAGCCACCTGGACGCGGCCTGGAATGGATCGGCTACATTTCTTATAGTGGGATCACCACATACAACCCCTCTCTGAAGAGTCGAGTGACCATTTCCAGAGACAACTCTAAAAATACACTGTATCTGCAGATGAATAGTCTGCGGGCCGAGGATACAGCTGTGTACTATTGTGCACGGTCTCTGGCCAGAACTACCGCTATGGACTATTGGGGGCAGGGAAGCCTGGTGACCGTCAGCTCCGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA] Amino Acid Sequence[spacer (SEQ ID NO: 87)][pLW085 without spacer (SEQ ID NO: 514)](SEQ ID NO: 472)[QGQSGQ][MMYCGGNEVLCGPRVGSSGGSGGSGGLSGRSDNHGGGSQTVVTQEPSLTVSPGGTVTLTCRSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGTPARFSGSLLGGKAALTLSGVQPEDEAEYYCALWYSNLWVFGGGTKLTVLGGGGSGGGGSGGGGSEVQLVESGGGLVQPGGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKNTAYLQMNNLKTEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSSGGGGSQVQLQESGPGLVRPSQTLSLTCTVSGYSITSDHAWSWVRQPPGRGLEWIGYISYSGITTYNPSLKSRVTISRDNSKNTLYLQMNSLRAEDTAVYYCARSLARTTAMDYWGQGSLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]*pLW079: LC IL6R 4792 Nsub Nucleotide Sequence[spacer (SEQ ID NO: 507)][pLW079 without spacer (SEQ ID NO: 527)](SEQ ID NO: 473)[CAAGGCCAGTCTGGCCAG][TATGGGTCCTGCAGTTGGAACTATGTACACATATTCATGGATTGCGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGCTCAGGTGGAGGCTCGGGCGGTGGGAGCGGCGGTTCTGACATCCAGATGACTCAGTCTCCTAGCTCCCTGTCCGCCTCTGTGGGGGACCGAGTCACCATCACATGCAGAGCCAGCCAGGATATTTCTAGTTACCTGAACTGGTATCAGCAGAAGCCCGGAAAAGCACCTAAGCTGCTGATCTACTATACCTCCAGGCTGCACTCTGGCGTGCCCAGTCGGTTCAGTGGCTCAGGGAGCGGAACCGACTTCACTTTTACCATCTCAAGCCTGCAGCCAGAGGATATTGCCACATACTATTGTCAGCAGGGCAATACACTGCCCTACACTTTTGGCCAGGGGACCAAGGTGGAAATCAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG] Amino Acid Sequence[spacer (SEQ ID NO: 87)][pLW079 without spacer (SEQ ID NO: 528)](SEQ ID NO: 474)[QGQSGQ][YGSCSWNYVHIFMDCGSSGGSGGSGGSGGGSGGGSGGSDIQMTQSPSSLSASVGDRVTITCRASQDISSYLNWYQQKPGKAPKLLIYYTSRLHSGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQGNTLPYTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC]* CI030: IL6RN297Q-15865 Nsub-CD3LvHv-H-NpLW087: HC IL6RN297Q-15865_Nsub-CD3LvHv-H-N Nucleotide Sequence[spacer (SEQ ID NO: 507)][pLW087 without spacer (SEQ ID NO: 529)](SEQ ID NO: 477)[CAAGGCCAGTCTGGCCAA][ATGATGTATTGCGGTGGGAATGAGGTGTTGTGCGGGCCGCGGGTTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTGGTGGAGGCTCGGGCGGTGGGAGCGGCGGCGGTTCTCAGACCGTGGTCACACAGGAGCCCTCACTGACAGTGAGCCCTGGCGGGACCGTCACACTGACTTGTCGCAGTTCAACTGGCGCCGTGACTACCAGCAATTACGCTAACTGGGTCCAGCAGAAACCAGGACAGGCACCACGAGGACTGATCGGAGGAACTAATAAGAGAGCACCAGGAACCCCTGCAAGGTTCTCCGGATCTCTGCTGGGGGGAAAAGCCGCTCTGACACTGAGCGGCGTGCAGCCTGAGGACGAAGCTGAGTACTATTGCGCACTGTGGTACTCCAACCTGTGGGTGTTTGGCGGGGGAACTAAGCTGACCGTCCTGGGAGGAGGAGGAAGCGGAGGAGGAGGGAGCGGAGGAGGAGGATCCGAAGTGCAGCTGGTCGAGAGCGGAGGAGGACTGGTGCAGCCAGGAGGATCCCTGAAGCTGTCTTGTGCAGCCAGTGGCTTCACCTTCAACACTTACGCAATGAACTGGGTGCGGCAGGCACCTGGGAAGGGACTGGAATGGGTCGCCCGGATCAGATCTAAATACAATAACTATGCCACCTACTATGCTGACAGTGTGAAGGATAGGTTCACCATTTCACGCGACGATAGCAAAAACACAGCTTATCTGCAGATGAATAACCTGAAGACCGAGGATACAGCAGTGTACTATTGCGTCAGACACGGCAATTTCGGGAACTCTTACGTGAGTTGGTTTGCCTATTGGGGACAGGGGACACTGGTCACCGTCTCCTCAGGAGGTGGTGGATCCCAGGTGCAGCTGCAGGAGTCCGGACCAGGACTGGTCCGGCCCTCACAGACTCTGAGCCTGACATGCACTGTGTCAGGCTACAGCATCACCTCCGATCACGCCTGGAGCTGGGTCAGGCAGCCACCTGGACGCGGCCTGGAATGGATCGGCTACATTTCTTATAGTGGGATCACCACATACAACCCCTCTCTGAAGAGTCGAGTGACCATTTCCAGAGACAACTCTAAAAATACACTGTATCTGCAGATGAATAGTCTGCGGGCCGAGGATACAGCTGTGTACTATTGTGCACGGTCTCTGGCCAGAACTACCGCTATGGACTATTGGGGGCAGGGAAGCCTGGTGACCGTCAGCTCCGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA] Amino Acid Sequence[spacer (SEQ ID NO: 87)][pLW087 without spacer (SEQ ID NO: 530)](SEQ ID NO: 478)[QGQSGQ][MMYCGGNEVLCGPRVGSSGGSGGSGGGGGSGGGSGGGSQTVVTQEPSLTVSPGGTVTLTCRSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGTPARFSGSLLGGKAALTLSGVQPEDEAEYYCALWYSNLWVFGGGTKLTVLGGGGSGGGGSGGGGSEVQLVESGGGLVQPGGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKNTAYLQMNNLKTEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSSGGGGSQVQLQESGPGLVRPSQTLSLTCTVSGYSITSDHAWSWVRQPPGRGLEWIGYISYSGITTYNPSLKSRVTISRDNSKNTLYLQMNSLRAEDTAVYYCARSLARTTAMDYWGQGSLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]*pLW077: LC IL6R Nucleotide Sequence (SEQ ID NO: 467)GACATCCAGATGACTCAGTCTCCTAGCTCCCTGTCCGCCTCTGTGGGGGACCGAGTCACCATCACATGCAGAGCCAGCCAGGATATTTCTAGTTACCTGAACTGGTATCAGCAGAAGCCCGGAAAAGCACCTAAGCTGCTGATCTACTATACCTCCAGGCTGCACTCTGGCGTGCCCAGTCGGTTCAGTGGCTCAGGGAGCGGAACCGACTTCACTTTTACCATCTCAAGCCTGCAGCCAGAGGATATTGCCACATACTATTGTCAGCAGGGCAATACACTGCCCTACACTTTTGGCCAGGGGACCAAGGTGGAAATCAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG  Amino Acid Sequence (SEQ ID NO: 468)DIQMTQSPSSLSASVGDRVTITCRASQDISSYLNWYQQKPGKAPKLLIYYTSRLHSGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQGNTLPYTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC*CI031: IL6R 4792 Nsub N297Q-15865 Nsub-CD3LvHv-H-NpLW087: HC IL6RN297Q-15865_Nsub-CD3LvHv-H-N Nucleotide Sequence[spacer (SEQ ID NO: 507)][pLW087 without spacer (SEQ ID NO: 529)](SEQ ID NO: 477)[CAAGGCCAGTCTGGCCAA][ATGATGTATTGCGGTGGGAATGAGGTGTTGTGCGGGCCGCGGGTTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTGGTGGAGGCTCGGGCGGTGGGAGCGGCGGCGGTTCTCAGACCGTGGTCACACAGGAGCCCTCACTGACAGTGAGCCCTGGCGGGACCGTCACACTGACTTGTCGCAGTTCAACTGGCGCCGTGACTACCAGCAATTACGCTAACTGGGTCCAGCAGAAACCAGGACAGGCACCACGAGGACTGATCGGAGGAACTAATAAGAGAGCACCAGGAACCCCTGCAAGGTTCTCCGGATCTCTGCTGGGGGGAAAAGCCGCTCTGACACTGAGCGGCGTGCAGCCTGAGGACGAAGCTGAGTACTATTGCGCACTGTGGTACTCCAACCTGTGGGTGTTTGGCGGGGGAACTAAGCTGACCGTCCTGGGAGGAGGAGGAAGCGGAGGAGGAGGGAGCGGAGGAGGAGGATCCGAAGTGCAGCTGGTCGAGAGCGGAGGAGGACTGGTGCAGCCAGGAGGATCCCTGAAGCTGTCTTGTGCAGCCAGTGGCTTCACCTTCAACACTTACGCAATGAACTGGGTGCGGCAGGCACCTGGGAAGGGACTGGAATGGGTCGCCCGGATCAGATCTAAATACAATAACTATGCCACCTACTATGCTGACAGTGTGAAGGATAGGTTCACCATTTCACGCGACGATAGCAAAAACACAGCTTATCTGCAGATGAATAACCTGAAGACCGAGGATACAGCAGTGTACTATTGCGTCAGACACGGCAATTTCGGGAACTCTTACGTGAGTTGGTTTGCCTATTGGGGACAGGGGACACTGGTCACCGTCTCCTCAGGAGGTGGTGGATCCCAGGTGCAGCTGCAGGAGTCCGGACCAGGACTGGTCCGGCCCTCACAGACTCTGAGCCTGACATGCACTGTGTCAGGCTACAGCATCACCTCCGATCACGCCTGGAGCTGGGTCAGGCAGCCACCTGGACGCGGCCTGGAATGGATCGGCTACATTTCTTATAGTGGGATCACCACATACAACCCCTCTCTGAAGAGTCGAGTGACCATTTCCAGAGACAACTCTAAAAATACACTGTATCTGCAGATGAATAGTCTGCGGGCCGAGGATACAGCTGTGTACTATTGTGCACGGTCTCTGGCCAGAACTACCGCTATGGACTATTGGGGGCAGGGAAGCCTGGTGACCGTCAGCTCCGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA] Amino Acid Sequence[spacer (SEQ ID NO: 87)][pLW087 without spacer (SEQ ID NO: 530)](SEQ ID NO: 478)[QGQSGQ][MMYCGGNEVLCGPRVGSSGGSGGSGGGGGSGGGSGGGSQTVVTQEPSLTVSPGGTVTLTCRSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGTPARFSGSLLGGKAALTLSGVQPEDEAEYYCALWYSNLWVFGGGTKLTVLGGGGSGGGGSGGGGSEVQLVESGGGLVQPGGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKNTAYLQMNNLKTEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSSGGGGSQVQLQESGPGLVRPSQTLSLTCTVSGYSITSDHAWSWVRQPPGRGLEWIGYISYSGITTYNPSLKSRVTISRDNSKNTLYLQMNSLRAEDTAVYYCARSLARTTAMDYWGQGSLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]* pLW079: LC IL6R 4792 Nsub Nucleotide Sequence[spacer (SEQ ID NO: 507)][pLW079 without spacer (SEQ ID NO: 527)](SEQ ID NO: 473)[CAAGGCCAGTCTGGCCAG][TATGGGTCCTGCAGTTGGAACTATGTACACATATTCATGGATTGCGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGCTCAGGTGGAGGCTCGGGCGGTGGGAGCGGCGGTTCTGACATCCAGATGACTCAGTCTCCTAGCTCCCTGTCCGCCTCTGTGGGGGACCGAGTCACCATCACATGCAGAGCCAGCCAGGATATTTCTAGTTACCTGAACTGGTATCAGCAGAAGCCCGGAAAAGCACCTAAGCTGCTGATCTACTATACCTCCAGGCTGCACTCTGGCGTGCCCAGTCGGTTCAGTGGCTCAGGGAGCGGAACCGACTTCACTTTTACCATCTCAAGCCTGCAGCCAGAGGATATTGCCACATACTATTGTCAGCAGGGCAATACACTGCCCTACACTTTTGGCCAGGGGACCAAGGTGGAAATCAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG] Amino Acid Sequence[spacer (SEQ ID NO: 87)][pLW079 without spacer (SEQ ID NO: 528)](SEQ ID NO: 474)[QGQSGQ][YGSCSWNYVHIFMDCGSSGGSGGSGGSGGGSGGGSGGSDIQMTQSPSSLSASVGDRVTITCRASQDISSYLNWYQQKPGKAPKLLIYYTSRLHSGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQGNTLPYTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC]* CI032: 4792-1204-IL6RN297Q-15865 Nsub-CD3LvHv-H-NpLW087: HC IL6RN297Q-15865_Nsub-CD3LvHv-H-N Nucleotide Sequence[spacer (SEQ ID NO: 507)][pLW087 without spacer (SEQ ID NO: 529)](SEQ ID NO: 477)[CAAGGCCAGTCTGGCCAA][ATGATGTATTGCGGTGGGAATGAGGTGTTGTGCGGGCCGCGGGTTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTGGTGGAGGCTCGGGCGGTGGGAGCGGCGGCGGTTCTCAGACCGTGGTCACACAGGAGCCCTCACTGACAGTGAGCCCTGGCGGGACCGTCACACTGACTTGTCGCAGTTCAACTGGCGCCGTGACTACCAGCAATTACGCTAACTGGGTCCAGCAGAAACCAGGACAGGCACCACGAGGACTGATCGGAGGAACTAATAAGAGAGCACCAGGAACCCCTGCAAGGTTCTCCGGATCTCTGCTGGGGGGAAAAGCCGCTCTGACACTGAGCGGCGTGCAGCCTGAGGACGAAGCTGAGTACTATTGCGCACTGTGGTACTCCAACCTGTGGGTGTTTGGCGGGGGAACTAAGCTGACCGTCCTGGGAGGAGGAGGAAGCGGAGGAGGAGGGAGCGGAGGAGGAGGATCCGAAGTGCAGCTGGTCGAGAGCGGAGGAGGACTGGTGCAGCCAGGAGGATCCCTGAAGCTGTCTTGTGCAGCCAGTGGCTTCACCTTCAACACTTACGCAATGAACTGGGTGCGGCAGGCACCTGGGAAGGGACTGGAATGGGTCGCCCGGATCAGATCTAAATACAATAACTATGCCACCTACTATGCTGACAGTGTGAAGGATAGGTTCACCATTTCACGCGACGATAGCAAAAACACAGCTTATCTGCAGATGAATAACCTGAAGACCGAGGATACAGCAGTGTACTATTGCGTCAGACACGGCAATTTCGGGAACTCTTACGTGAGTTGGTTTGCCTATTGGGGACAGGGGACACTGGTCACCGTCTCCTCAGGAGGTGGTGGATCCCAGGTGCAGCTGCAGGAGTCCGGACCAGGACTGGTCCGGCCCTCACAGACTCTGAGCCTGACATGCACTGTGTCAGGCTACAGCATCACCTCCGATCACGCCTGGAGCTGGGTCAGGCAGCCACCTGGACGCGGCCTGGAATGGATCGGCTACATTTCTTATAGTGGGATCACCACATACAACCCCTCTCTGAAGAGTCGAGTGACCATTTCCAGAGACAACTCTAAAAATACACTGTATCTGCAGATGAATAGTCTGCGGGCCGAGGATACAGCTGTGTACTATTGTGCACGGTCTCTGGCCAGAACTACCGCTATGGACTATTGGGGGCAGGGAAGCCTGGTGACCGTCAGCTCCGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA] Amino Acid Sequence[spacer (SEQ ID NO: 87)][pLW087 without spacer (SEQ ID NO: 530)](SEQ ID NO: 478)[QGQSGQ][MMYCGGNEVLCGPRVGSSGGSGGSGGGGGSGGGSGGGSQTVVTQEPSLTVSPGGTVTLTCRSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGTPARFSGSLLGGKAALTLSGVQPEDEAEYYCALWYSNLWVFGGGTKLTVLGGGGSGGGGSGGGGSEVQLVESGGGLVQPGGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKNTAYLQMNNLKTEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSSGGGGSQVQLQESGPGLVRPSQTLSLTCTVSGYSITSDHAWSWVRQPPGRGLEWIGYISYSGITTYNPSLKSRVTISRDNSKNTLYLQMNSLRAEDTAVYYCARSLARTTAMDYWGQGSLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]* pLW080: LC IL6R 4792 1204 Nucleotide Sequence[spacer (SEQ ID NO: 507)][pLW080 without spacer (SEQ ID NO: 515)](SEQ ID NO: 473)[CAAGGCCAGTCTGGCCAG][TATGGGTCCTGCAGTTGGAACTATGTACACATATTCATGGATTGCGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTGACATCCAGATGACTCAGTCTCCTAGCTCCCTGTCCGCCTCTGTGGGGGACCGAGTCACCATCACATGCAGAGCCAGCCAGGATATTTCTAGTTACCTGAACTGGTATCAGCAGAAGCCCGGAAAAGCACCTAAGCTGCTGATCTACTATACCTCCAGGCTGCACTCTGGCGTGCCCAGTCGGTTCAGTGGCTCAGGGAGCGGAACCGACTTCACTTTTACCATCTCAAGCCTGCAGCCAGAGGATATTGCCACATACTATTGTCAGCAGGGCAATACACTGCCCTACACTTTTGGCCAGGGGACCAAGGTGGAAATCAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG] Amino Acid Sequence[spacer (SEQ ID NO: 87)][pLW080 without spacer (SEQ ID NO: 516)](SEQ ID NO: 474)[QGQSGQ][YGSCSWNYVHIFMDCGSSGGSGGSGGLSGRSDNHGGGSDIQMTQSPSSLSASVGDRVTITCRASQDISSYLNWYQQKPGKAPKLLIYYTSRLHSGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQGNTLPYTFGQGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC]*  CI048: Activated CI011HC C225v5N297Q-*CD3LvHv-H-N Nucleotide Sequence (SEQ ID NO: 451)CAAGGCCAGTCTGGCCAAATGATGTATTGCGGTGGGAATGAGGTGTTGTGCGGGCCGCGGGTTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTCTGAGCGGCCGTTCCGATAATCATGGCGGCGGTTCTCAGACCGTGGTCACACAGGAGCCCTCACTGACAGTGAGCCCTGGCGGGACCGTCACACTGACTTGTCGCAGTTCAACTGGCGCCGTGACTACCAGCAATTACGCTAACTGGGTCCAGCAGAAACCAGGACAGGCACCACGAGGACTGATCGGAGGAACTAATAAGAGAGCACCAGGAACCCCTGCAAGGTTCTCCGGATCTCTGCTGGGGGGAAAAGCCGCTCTGACACTGAGCGGCGTGCAGCCTGAGGACGAAGCTGAGTACTATTGCGCACTGTGGTACTCCAACCTGTGGGTGTTTGGCGGGGGAACTAAGCTGACCGTCCTGGGAGGAGGAGGAAGCGGAGGAGGAGGGAGCGGAGGAGGAGGATCCGAAGTGCAGCTGGTCGAGAGCGGAGGAGGACTGGTGCAGCCAGGAGGATCCCTGAAGCTGTCTTGTGCAGCCAGTGGCTTCACCTTCAACACTTACGCAATGAACTGGGTGCGGCAGGCACCTGGGAAGGGACTGGAATGGGTCGCCCGGATCAGATCTAAATACAATAACTATGCCACCTACTATGCTGACAGTGTGAAGGATAGGTTCACCATTTCACGCGACGATAGCAAAAACACAGCTTATCTGCAGATGAATAACCTGAAGACCGAGGATACAGCAGTGTACTATTGCGTCAGACACGGCAATTTCGGGAACTCTTACGTGAGTTGGTTTGCCTATTGGGGACAGGGGACACTGGTCACCGTCTCCTCAGGAGGTGGTGGATCCCAGGTGCAGCTGAAACAGAGCGGCCCGGGCCTGGTGCAGCCGAGCCAGAGCCTGAGCATTACCTGCACCGTGAGCGGCTTTAGCCTGACCAACTATGGCGTGCATTGGGTGCGCCAGAGCCCGGGCAAAGGCCTGGAATGGCTGGGCGTGATTTGGAGCGGCGGCAACACCGATTATAACACCCCGTTTACCAGCCGCCTGAGCATTAACAAAGATAACAGCAAAAGCCAGGTGTTTTTTAAAATGAACAGCCTGCAAAGCCAGGATACCGCGATTTATTATTGCGCGCGCGCGCTGACCTATTATGATTATGAATTTGCGTATTGGGGCCAGGGCACCCTGGTGACCGTGAGCGCGGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA  Amino Acid Sequence (SEQ ID NO: 452)SDNHGGGSQTVVTQEPSLTVSPGGTVTLTCRSSTGAVTTSNYANWVQQKPGQAPRGLIGGTNKRAPGTPARFSGSLLGGKAALTLSGVQPEDEAEYYCALWYSNLWVFGGGTKLTVLGGGGSGGGGSGGGGSEVQLVESGGGLVQPGGSLKLSCAASGFTFNTYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKNTAYLQMNNLKTEDTAVYYCVRHGNFGNSYVSWFAYWGQGTLVTVSSGGGGSQVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSQDTAIYYCARALTYYDYEFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK Activated Light Chain Nucleotide Sequence(SEQ ID NO: 483)TCCGATAATCATGGCAGTAGCGGTACCCAGATCTTGCTGACCCAGAGCCCGGTGATTCTGAGCGTGAGCCCGGGCGAACGTGTGAGCTTTAGCTGCCGCGCGAGCCAGAGCATTGGCACCAACATTCATTGGTATCAGCAGCGCACCAACGGCAGCCCGCGCCTGCTGATTAAATATGCGAGCGAAAGCATTAGCGGCATTCCGAGCCGCTTTAGCGGCAGCGGCAGCGGCACCGATTTTACCCTGAGCATTAACAGCGTGGAAAGCGAAGATATTGCGGATTATTATTGCCAGCAGAACAACAACTGGCCGACCACCTTTGGCGCGGGCACCAAACTGGAACTGAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG Amino Acid Sequence (SEQ ID NO: 484)SDNHGSSGTQILLTQSPVILSVSPGERVSFSCRASQSIGTNIHWYQQRTNGSPRLLIKYASESISGIPSRFSGSGSGTDFTLSINSVESEDIADYYCQQNNNWPTTFGAGTKLELKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC CI052: C225v5N297Q-mCD3-H-N pLW100: HC C225v5N297Q-mCD3-H-NNucleotide Sequence (SEQ ID NO: 491)GAAGTGCAGCTGGTGGAATCTGGGGGCGGACTGGTGCAGCCTGGCAAGTCTCTGAAGCTGAGCTGCGAGGCCAGCGGCTTCACCTTTAGCGGCTACGGCATGCACTGGGTGCGCCAGGCACCTGGCAGAGGCCTGGAAAGCGTGGCCTACATCACCAGCAGCAGCATCAACATTAAGTACGCCGACGCCGTGAAGGGCCGGTTCACCGTGTCCAGAGACAACGCCAAGAACCTGCTGTTCCTGCAGATGAACATCCTGAAGTCCGAGGACACCGCCATGTACTACTGCGCCAGATTCGACTGGGACAAGAACTACTGGGGCCAGGGCACAATGGTCACAGTGTCCAGCGGTGGAGGTGGTAGTGGTGGAGGAAGTGGAGGTTCAGGAGGTGGAAGCGGTGGTGGTGGTAGTGACATCCAGATGACCCAGAGCCCCAGCAGCCTGCCTGCCTCTCTGGGCGATAGAGTGACCATCAACTGCCAGGCCAGCCAGGACATCAGCAACTACCTGAACTGGTATCAGCAGAAGCCCGGCAAGGCCCCCAAGCTGCTGATCTACTACACCAACAAGCTGGCCGACGGTGTGCCCAGCAGATTCAGCGGCAGCGGTAGCGGCAGAGACAGCAGCTTCACCATCAGCTCCCTGGAATCCGAGGATATCGGCAGCTACTACTGCCAGCAGTACTACAACTACCCCTGGACCTTCGGCCCTGGCACCAAGCTGGAAATCAAAAGAGGAGGTGGTGGATCCCAGGTGCAGCTGAAACAGAGCGGCCCGGGCCTGGTGCAGCCGAGCCAGAGCCTGAGCATTACCTGCACCGTGAGCGGCTTTAGCCTGACCAACTATGGCGTGCATTGGGTGCGCCAGAGCCCGGGCAAAGGCCTGGAATGGCTGGGCGTGATTTGGAGCGGCGGCAACACCGATTATAACACCCCGTTTACCAGCCGCCTGAGCATTAACAAAGATAACAGCAAAAGCCAGGTGTTTTTTAAAATGAACAGCCTGCAAAGCCAGGATACCGCGATTTATTATTGCGCGCGCGCGCTGACCTATTATGATTATGAATTTGCGTATTGGGGCCAGGGCACCCTGGTGACCGTGAGCGCGGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA  Amino Acid Sequence (SEQ ID NO: 492)EVQLVESGGGLVQPGKSLKLSCEASGFTFSGYGMHWVRQAPGRGLESVAYITSSSINIKYADAVKGRFTVSRDNAKNLLFLQMNILKSEDTAMYYCARFDWDKNYWGQGTMVTVSSGGGGSGGGSGGSGGGSGGGGSDIQMTQSPSSLPASLGDRVTINCQASQDISNYLNWYQQKPGKAPKLLIYYTNKLADGVPSRFSGSGSGRDSSFTISSLESEDIGSYYCQQYYNYPWTFGPGTKLEIKRGGGGSQVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSQDTAIYYCARALTYYDYEFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK*  OPP021: LC C225 Nucleotide Sequence (SEQ ID NO: 457)GATATCTTGCTGACCCAGAGCCCGGTGATTCTGAGCGTGAGCCCGGGCGAACGTGTGAGCTTTAGCTGCCGCGCGAGCCAGAGCATTGGCACCAACATTCATTGGTATCAGCAGCGCACCAACGGCAGCCCGCGCCTGCTGATTAAATATGCGAGCGAAAGCATTAGCGGCATTCCGAGCCGCTTTAGCGGCAGCGGCAGCGGCACCGATTTTACCCTGAGCATTAACAGCGTGGAAAGCGAAGATATTGCGGATTATTATTGCCAGCAGAACAACAACTGGCCGACCACCTTTGGCGCGGGCACCAAACTGGAAATCAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG  Amino Acid Sequence (SEQ ID NO: 458)DILLTQSPVILSVSPGERVSFSCRASQSIGTNIHWYQQRTNGSPRLLIKYASESISGIPSRFSGSGSGTDFTLSINSVESEDIADYYCQQNNNWPTTFGAGTKLEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC* CI053: 3954-1204-C225v5N297Q-MC01-2001-mCD3-H-NpLW117: HC C225v5N297Q-MC01-2001-mCD3-H-N Nucleotide Sequence[spacer (SEQ ID NO: 507)][pLW117 without spacer (SEQ ID NO: 531)](SEQ ID NO: 493)[CAAGGCCAGTCTGGCCAA][TTGCATCCTATGTGCCATCCTGAGGGTCTGTGCAAGTTTACTCCTGGAGGTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTATTAGCAGTGGTCTGTTAAGCGGTCGTAGCGATAATCATGGCGGCGGTTCTGAAGTGCAGCTGGTGGAATCTGGGGGCGGACTGGTGCAGCCTGGCAAGTCTCTGAAGCTGAGCTGCGAGGCCAGCGGCTTCACCTTTAGCGGCTACGGCATGCACTGGGTGCGCCAGGCACCTGGCAGAGGCCTGGAAAGCGTGGCCTACATCACCAGCAGCAGCATCAACATTAAGTACGCCGACGCCGTGAAGGGCCGGTTCACCGTGTCCAGAGACAACGCCAAGAACCTGCTGTTCCTGCAGATGAACATCCTGAAGTCCGAGGACACCGCCATGTACTACTGCGCCAGATTCGACTGGGACAAGAACTACTGGGGCCAGGGCACAATGGTCACAGTGTCCAGCGGTGGAGGTGGTAGTGGTGGAGGAAGTGGAGGTTCAGGAGGTGGAAGCGGTGGTGGTGGTAGTGACATCCAGATGACCCAGAGCCCCAGCAGCCTGCCTGCCTCTCTGGGCGATAGAGTGACCATCAACTGCCAGGCCAGCCAGGACATCAGCAACTACCTGAACTGGTATCAGCAGAAGCCCGGCAAGGCCCCCAAGCTGCTGATCTACTACACCAACAAGCTGGCCGACGGTGTGCCCAGCAGATTCAGCGGCAGCGGTAGCGGCAGAGACAGCAGCTTCACCATCAGCTCCCTGGAATCCGAGGATATCGGCAGCTACTACTGCCAGCAGTACTACAACTACCCCTGGACCTTCGGCCCTGGCACCAAGCTGGAAATCAAAAGAGGAGGTGGTGGATCCCAGGTGCAGCTGAAACAGAGCGGCCCGGGCCTGGTGCAGCCGAGCCAGAGCCTGAGCATTACCTGCACCGTGAGCGGCTTTAGCCTGACCAACTATGGCGTGCATTGGGTGCGCCAGAGCCCGGGCAAAGGCCTGGAATGGCTGGGCGTGATTTGGAGCGGCGGCAACACCGATTATAACACCCCGTTTACCAGCCGCCTGAGCATTAACAAAGATAACAGCAAAAGCCAGGTGTTTTTTAAAATGAACAGCCTGCAAAGCCAGGATACCGCGATTTATTATTGCGCGCGCGCGCTGACCTATTATGATTATGAATTTGCGTATTGGGGCCAGGGCACCCTGGTGACCGTGAGCGCGGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA] Amino Acid Sequence[spacer (SEQ ID NO: 87)][pLW117 without spacer (SEQ ID NO: 532)](SEQ ID NO: 494)[QGQSGQ][LHPMCHPEGLCKFTPGGGSSGGSGGSGGISSGLLSGRSDNHGGGSEVQLVESGGGLVQPGKSLKLSCEASGFTFSGYGMHWVRQAPGRGLESVAYITSSSINIKYADAVKGRFTVSRDNAKNLLFLQMNILKSEDTAMYYCARFDWDKNYWGQGTMVTVSSGGGGSGGGSGGSGGGSGGGGSDIQMTQSPSSLPASLGDRVTINCQASQDISNYLNWYQQKPGKAPKLLIYYTNKLADGVPSRFSGSGSGRDSSFTISSLESEDIGSYYCQQYYNYPWTFGPGTKLEIKRGGGGSQVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSQDTAIYYCARALTYYDYEFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]*OPP022: LC C225 3954-1204 Nucleotide Sequence[spacer (SEQ ID NO: 507)][OPP022 without spacer (SEQ ID NO: 509)](SEQ ID NO: 449)[CAAGGCCAGTCTGGCCAG][TGCATCTCACCTCGTGGTTGTCCGGACGGCCCATACGTCATGTACGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGATCCGGTCTGAGCGGCCGTTCCGATAATCATGGCAGTAGCGGTACCCAGATCTTGCTGACCCAGAGCCCGGTGATTCTGAGCGTGAGCCCGGGCGAACGTGTGAGCTTTAGCTGCCGCGCGAGCCAGAGCATTGGCACCAACATTCATTGGTATCAGCAGCGCACCAACGGCAGCCCGCGCCTGCTGATTAAATATGCGAGCGAAAGCATTAGCGGCATTCCGAGCCGCTTTAGCGGCAGCGGCAGCGGCACCGATTTTACCCTGAGCATTAACAGCGTGGAAAGCGAAGATATTGCGGATTATTATTGCCAGCAGAACAACAACTGGCCGACCACCTTTGGCGCGGGCACCAAACTGGAACTGAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG] Amino Acid Sequence[spacer (SEQ ID NO: 87)][OPP022 without spacer (SEQ ID NO: 508)](SEQ ID NO: 450)[QGQSGQ][CISPRGCPDGPYVMYGSSGGSGGSGGSGLSGRSDNHGSSGTQILLTQSPVILSVSPGERVSFSCRASQSIGTNIHWYQQRTNGSPRLLIKYASESISGIPSRFSGSGSGTDFTLSINSVESEDIADYYCQQNNNWPTTFGAGTKLELKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC]* CI054: 3954-1204-C225v5N297Q-MCO2-2001-mCD3-H-NpLW118: HC C225v5N297Q-MCO2-2001-mCD3-H-N Nucleotide Sequence[spacer (SEQ ID NO: 521)][pLW118 without spacer (SEQ ID NO: 533)](SEQ ID NO: 495)[CAAGGCCAGTCTGGCCAAGGT][GCTTGCTCTGATATGGTTTATTGGGGTTCGTGCAGTTGGTTGGGAGGTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTATTAGCAGTGGTCTGTTAAGCGGTCGTAGCGATAATCATGGCGGCGGTTCTGAAGTGCAGCTGGTGGAATCTGGGGGCGGACTGGTGCAGCCTGGCAAGTCTCTGAAGCTGAGCTGCGAGGCCAGCGGCTTCACCTTTAGCGGCTACGGCATGCACTGGGTGCGCCAGGCACCTGGCAGAGGCCTGGAAAGCGTGGCCTACATCACCAGCAGCAGCATCAACATTAAGTACGCCGACGCCGTGAAGGGCCGGTTCACCGTGTCCAGAGACAACGCCAAGAACCTGCTGTTCCTGCAGATGAACATCCTGAAGTCCGAGGACACCGCCATGTACTACTGCGCCAGATTCGACTGGGACAAGAACTACTGGGGCCAGGGCACAATGGTCACAGTGTCCAGCGGTGGAGGTGGTAGTGGTGGAGGAAGTGGAGGTTCAGGAGGTGGAAGCGGTGGTGGTGGTAGTGACATCCAGATGACCCAGAGCCCCAGCAGCCTGCCTGCCTCTCTGGGCGATAGAGTGACCATCAACTGCCAGGCCAGCCAGGACATCAGCAACTACCTGAACTGGTATCAGCAGAAGCCCGGCAAGGCCCCCAAGCTGCTGATCTACTACACCAACAAGCTGGCCGACGGTGTGCCCAGCAGATTCAGCGGCAGCGGTAGCGGCAGAGACAGCAGCTTCACCATCAGCTCCCTGGAATCCGAGGATATCGGCAGCTACTACTGCCAGCAGTACTACAACTACCCCTGGACCTTCGGCCCTGGCACCAAGCTGGAAATCAAAAGAGGAGGTGGTGGATCCCAGGTGCAGCTGAAACAGAGCGGCCCGGGCCTGGTGCAGCCGAGCCAGAGCCTGAGCATTACCTGCACCGTGAGCGGCTTTAGCCTGACCAACTATGGCGTGCATTGGGTGCGCCAGAGCCCGGGCAAAGGCCTGGAATGGCTGGGCGTGATTTGGAGCGGCGGCAACACCGATTATAACACCCCGTTTACCAGCCGCCTGAGCATTAACAAAGATAACAGCAAAAGCCAGGTGTTTTTTAAAATGAACAGCCTGCAAAGCCAGGATACCGCGATTTATTATTGCGCGCGCGCGCTGACCTATTATGATTATGAATTTGCGTATTGGGGCCAGGGCACCCTGGTGACCGTGAGCGCGGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA] Amino Acid Sequence[spacer (SEQ ID NO: 407)][pLW118 without spacer (SEQ ID NO: 534)](SEQ ID NO: 496)[QGQSGQG][ACSDMVYWGSCSWLGGGSSGGSGGSGGISSGLLSGRSDNHGGGSEVQLVESGGGLVQPGKSLKLSCEASGFTFSGYGMHWVRQAPGRGLESVAYITSSSINIKYADAVKGRFTVSRDNAKNLLFLQMNILKSEDTAMYYCARFDWDKNYWGQGTMVTVSSGGGGSGGGSGGSGGGSGGGGSDIQMTQSPSSLPASLGDRVTINCQASQDISNYLNWYQQKPGKAPKLLIYYTNKLADGVPSRFSGSGSGRDSSFTISSLESEDIGSYYCQQYYNYPWTFGPGTKLEIKRGGGGSQVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSQDTAIYYCARALTYYDYEFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]*OPP022: LC C225 3954-1204 Nucleotide Sequence[spacer (SEQ ID NO: 507)][OPP022 without spacer (SEQ ID NO: 509)](SEQ ID NO: 449)[CAAGGCCAGTCTGGCCAG][TGCATCTCACCTCGTGGTTGTCCGGACGGCCCATACGTCATGTACGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGATCCGGTCTGAGCGGCCGTTCCGATAATCATGGCAGTAGCGGTACCCAGATCTTGCTGACCCAGAGCCCGGTGATTCTGAGCGTGAGCCCGGGCGAACGTGTGAGCTTTAGCTGCCGCGCGAGCCAGAGCATTGGCACCAACATTCATTGGTATCAGCAGCGCACCAACGGCAGCCCGCGCCTGCTGATTAAATATGCGAGCGAAAGCATTAGCGGCATTCCGAGCCGCTTTAGCGGCAGCGGCAGCGGCACCGATTTTACCCTGAGCATTAACAGCGTGGAAAGCGAAGATATTGCGGATTATTATTGCCAGCAGAACAACAACTGGCCGACCACCTTTGGCGCGGGCACCAAACTGGAACTGAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG] Amino Acid Sequence[spacer (SEQ ID NO: 87)][OPP022 without spacer (SEQ ID NO: 508)](SEQ ID NO: 450)[QGQSGQ][CISPRGCPDGPYVMYGSSGGSGGSGGSGLSGRSDNHGSSGTQILLTQSPVILSVSPGERVSFSCRASQSIGTNIHWYQQRTNGSPRLLIKYASESISGIPSRFSGSGSGTDFTLSINSVESEDIADYYCQQNNNWPTTFGAGTKLELKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC]*CI055: 3954-1204-C225v5N297Q-MC03-2001-mCD3-H-NpLW119: HC C225v5N297Q-MC03-2001-mCD3-H-N Nucleotide Sequence[spacer (SEQ ID NO: 507)][pLW119 without spacer (SEQ ID NO: 535)](SEQ ID NO: 497)[CAAGGCCAGTCTGGCCAA][GATTGCATTGGATTGGATCATTATTTTCTTGGACCGTGCAGTTCTGGAGGTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTATTAGCAGTGGTCTGTTAAGCGGTCGTAGCGATAATCATGGCGGCGGTTCTGAAGTGCAGCTGGTGGAATCTGGGGGCGGACTGGTGCAGCCTGGCAAGTCTCTGAAGCTGAGCTGCGAGGCCAGCGGCTTCACCTTTAGCGGCTACGGCATGCACTGGGTGCGCCAGGCACCTGGCAGAGGCCTGGAAAGCGTGGCCTACATCACCAGCAGCAGCATCAACATTAAGTACGCCGACGCCGTGAAGGGCCGGTTCACCGTGTCCAGAGACAACGCCAAGAACCTGCTGTTCCTGCAGATGAACATCCTGAAGTCCGAGGACACCGCCATGTACTACTGCGCCAGATTCGACTGGGACAAGAACTACTGGGGCCAGGGCACAATGGTCACAGTGTCCAGCGGTGGAGGTGGTAGTGGTGGAGGAAGTGGAGGTTCAGGAGGTGGAAGCGGTGGTGGTGGTAGTGACATCCAGATGACCCAGAGCCCCAGCAGCCTGCCTGCCTCTCTGGGCGATAGAGTGACCATCAACTGCCAGGCCAGCCAGGACATCAGCAACTACCTGAACTGGTATCAGCAGAAGCCCGGCAAGGCCCCCAAGCTGCTGATCTACTACACCAACAAGCTGGCCGACGGTGTGCCCAGCAGATTCAGCGGCAGCGGTAGCGGCAGAGACAGCAGCTTCACCATCAGCTCCCTGGAATCCGAGGATATCGGCAGCTACTACTGCCAGCAGTACTACAACTACCCCTGGACCTTCGGCCCTGGCACCAAGCTGGAAATCAAAAGAGGAGGTGGTGGATCCCAGGTGCAGCTGAAACAGAGCGGCCCGGGCCTGGTGCAGCCGAGCCAGAGCCTGAGCATTACCTGCACCGTGAGCGGCTTTAGCCTGACCAACTATGGCGTGCATTGGGTGCGCCAGAGCCCGGGCAAAGGCCTGGAATGGCTGGGCGTGATTTGGAGCGGCGGCAACACCGATTATAACACCCCGTTTACCAGCCGCCTGAGCATTAACAAAGATAACAGCAAAAGCCAGGTGTTTTTTAAAATGAACAGCCTGCAAAGCCAGGATACCGCGATTTATTATTGCGCGCGCGCGCTGACCTATTATGATTATGAATTTGCGTATTGGGGCCAGGGCACCCTGGTGACCGTGAGCGCGGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA] Amino Acid Sequence[spacer (SEQ ID NO: 87)][pLW119 without spacer (SEQ ID NO: 536)](SEQ ID NO: 498)[QGQSGQ][DCIGLDHYFLGPCSSGGGSSGGSGGSGGISSGLLSGRSDNHGGGSEVQLVESGGGLVQPGKSLKLSCEASGFTFSGYGMHWVRQAPGRGLESVAYITSSSINIKYADAVKGRFTVSRDNAKNLLFLQMNILKSEDTAMYYCARFDWDKNYWGQGTMVTVSSGGGGSGGGSGGSGGGSGGGGSDIQMTQSPSSLPASLGDRVTINCQASQDISNYLNWYQQKPGKAPKLLIYYTNKLADGVPSRFSGSGSGRDSSFTISSLESEDIGSYYCQQYYNYPWTFGPGTKLEIKRGGGGSQVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSQDTAIYYCARALTYYDYEFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]*OPP022: LC C225 3954-1204 Nucleotide Sequence[spacer (SEQ ID NO: 507)][OPP022 without spacer (SEQ ID NO: 509)](SEQ ID NO: 449)[CAAGGCCAGTCTGGCCAG][TGCATCTCACCTCGTGGTTGTCCGGACGGCCCATACGTCATGTACGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGATCCGGTCTGAGCGGCCGTTCCGATAATCATGGCAGTAGCGGTACCCAGATCTTGCTGACCCAGAGCCCGGTGATTCTGAGCGTGAGCCCGGGCGAACGTGTGAGCTTTAGCTGCCGCGCGAGCCAGAGCATTGGCACCAACATTCATTGGTATCAGCAGCGCACCAACGGCAGCCCGCGCCTGCTGATTAAATATGCGAGCGAAAGCATTAGCGGCATTCCGAGCCGCTTTAGCGGCAGCGGCAGCGGCACCGATTTTACCCTGAGCATTAACAGCGTGGAAAGCGAAGATATTGCGGATTATTATTGCCAGCAGAACAACAACTGGCCGACCACCTTTGGCGCGGGCACCAAACTGGAACTGAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG] Amino Acid Sequence[spacer (SEQ ID NO: 87)][OPP022 without spacer (SEQ ID NO: 508)](SEQ ID NO: 450)[QGQSGQ][CISPRGCPDGPYVMYGSSGGSGGSGGSGLSGRSDNHGSSGTQILLTQSPVILSVSPGERVSFSCRASQSIGTNIHWYQQRTNGSPRLLIKYASESISGIPSRFSGSGSGTDFTLSINSVESEDIADYYCQQNNNWPTTFGAGTKLELKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC]*CI056: 3954-1204-C225v5N297Q-MC04-2001-mCD3-H-NpLW120: HC C225v5N297Q-MC04-2001-mCD3-H-N Nucleotide Sequence[spacer (SEQ ID NO: 507)][pLW120 without spacer (SEQ ID NO: 537)](SEQ ID NO: 499)[CAAGGCCAGTCTGGCCAA][TCTATGTGCACTGAGCAGCAGTGGATTGTGAATCATTGCATTAGTGGAGGTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTATTAGCAGTGGTCTGTTAAGCGGTCGTAGCGATAATCATGGCGGCGGTTCTGAAGTGCAGCTGGTGGAATCTGGGGGCGGACTGGTGCAGCCTGGCAAGTCTCTGAAGCTGAGCTGCGAGGCCAGCGGCTTCACCTTTAGCGGCTACGGCATGCACTGGGTGCGCCAGGCACCTGGCAGAGGCCTGGAAAGCGTGGCCTACATCACCAGCAGCAGCATCAACATTAAGTACGCCGACGCCGTGAAGGGCCGGTTCACCGTGTCCAGAGACAACGCCAAGAACCTGCTGTTCCTGCAGATGAACATCCTGAAGTCCGAGGACACCGCCATGTACTACTGCGCCAGATTCGACTGGGACAAGAACTACTGGGGCCAGGGCACAATGGTCACAGTGTCCAGCGGTGGAGGTGGTAGTGGTGGAGGAAGTGGAGGTTCAGGAGGTGGAAGCGGTGGTGGTGGTAGTGACATCCAGATGACCCAGAGCCCCAGCAGCCTGCCTGCCTCTCTGGGCGATAGAGTGACCATCAACTGCCAGGCCAGCCAGGACATCAGCAACTACCTGAACTGGTATCAGCAGAAGCCCGGCAAGGCCCCCAAGCTGCTGATCTACTACACCAACAAGCTGGCCGACGGTGTGCCCAGCAGATTCAGCGGCAGCGGTAGCGGCAGAGACAGCAGCTTCACCATCAGCTCCCTGGAATCCGAGGATATCGGCAGCTACTACTGCCAGCAGTACTACAACTACCCCTGGACCTTCGGCCCTGGCACCAAGCTGGAAATCAAAAGAGGAGGTGGTGGATCCCAGGTGCAGCTGAAACAGAGCGGCCCGGGCCTGGTGCAGCCGAGCCAGAGCCTGAGCATTACCTGCACCGTGAGCGGCTTTAGCCTGACCAACTATGGCGTGCATTGGGTGCGCCAGAGCCCGGGCAAAGGCCTGGAATGGCTGGGCGTGATTTGGAGCGGCGGCAACACCGATTATAACACCCCGTTTACCAGCCGCCTGAGCATTAACAAAGATAACAGCAAAAGCCAGGTGTTTTTTAAAATGAACAGCCTGCAAAGCCAGGATACCGCGATTTATTATTGCGCGCGCGCGCTGACCTATTATGATTATGAATTTGCGTATTGGGGCCAGGGCACCCTGGTGACCGTGAGCGCGGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA  Amino Acid Sequence[spacer (SEQ ID NO: 87)][pLW120 without spacer (SEQ ID NO: 538)](SEQ ID NO: 500)[QGQSGQ][SMCTEQQWIVNHCISGGGSSGGSGGSGGISSGLLSGRSDNHGGGSEVQLVESGGGLVQPGKSLKLSCEASGFTFSGYGMHWVRQAPGRGLESVAYITSSSINIKYADAVKGRFTVSRDNAKNLLFLQMNILKSEDTAMYYCARFDWDKNYWGQGTMVTVSSGGGGSGGGSGGSGGGSGGGGSDIQMTQSPSSLPASLGDRVTINCQASQDISNYLNWYQQKPGKAPKLLIYYTNKLADGVPSRFSGSGSGRDSSFTISSLESEDIGSYYCQQYYNYPWTFGPGTKLEIKRGGGGSQVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSQDTAIYYCARALTYYDYEFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]*OPP022: LC C225 3954-1204 Nucleotide Sequence[spacer (SEQ ID NO: 507)][OPP022 without spacer (SEQ ID NO: 509)](SEQ ID NO: 449)[CAAGGCCAGTCTGGCCAG][TGCATCTCACCTCGTGGTTGTCCGGACGGCCCATACGTCATGTACGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGATCCGGTCTGAGCGGCCGTTCCGATAATCATGGCAGTAGCGGTACCCAGATCTTGCTGACCCAGAGCCCGGTGATTCTGAGCGTGAGCCCGGGCGAACGTGTGAGCTTTAGCTGCCGCGCGAGCCAGAGCATTGGCACCAACATTCATTGGTATCAGCAGCGCACCAACGGCAGCCCGCGCCTGCTGATTAAATATGCGAGCGAAAGCATTAGCGGCATTCCGAGCCGCTTTAGCGGCAGCGGCAGCGGCACCGATTTTACCCTGAGCATTAACAGCGTGGAAAGCGAAGATATTGCGGATTATTATTGCCAGCAGAACAACAACTGGCCGACCACCTTTGGCGCGGGCACCAAACTGGAACTGAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG] Amino Acid Sequence[spacer (SEQ ID NO: 87)][OPP022 without spacer (SEQ ID NO: 508)](SEQ ID NO: 450)[QGQSGQ][CISPRGCPDGPYVMYGSSGGSGGSGGSGLSGRSDNHGSSGTQILLTQSPVILSVSPGERVSFSCRASQSIGTNIHWYQQRTNGSPRLLIKYASESISGIPSRFSGSGSGTDFTLSINSVESEDIADYYCQQNNNWPTTFGAGTKLELKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC]*CI049: 3954-1204-C225v5N297Q-MC05-2001-mCD3-H-NpLW121: HC C225v5N297Q-MC05-2001-mCD3-H-N Nucleotide Sequence[spacer (SEQ ID NO: 507)][pLW121 without spacer (SEQ ID NO: 539)](SEQ ID NO: 485)[CAAGGCCAGTCTGGCCAA][ATTCATCCGATGTGCCATCCTGAGGGTGTTTGCGTTGCTCTGGATGGAGGTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTATTAGCAGTGGTCTGTTAAGCGGTCGTAGCGATAATCATGGCGGCGGTTCTGAAGTGCAGCTGGTGGAATCTGGGGGCGGACTGGTGCAGCCTGGCAAGTCTCTGAAGCTGAGCTGCGAGGCCAGCGGCTTCACCTTTAGCGGCTACGGCATGCACTGGGTGCGCCAGGCACCTGGCAGAGGCCTGGAAAGCGTGGCCTACATCACCAGCAGCAGCATCAACATTAAGTACGCCGACGCCGTGAAGGGCCGGTTCACCGTGTCCAGAGACAACGCCAAGAACCTGCTGTTCCTGCAGATGAACATCCTGAAGTCCGAGGACACCGCCATGTACTACTGCGCCAGATTCGACTGGGACAAGAACTACTGGGGCCAGGGCACAATGGTCACAGTGTCCAGCGGTGGAGGTGGTAGTGGTGGAGGAAGTGGAGGTTCAGGAGGTGGAAGCGGTGGTGGTGGTAGTGACATCCAGATGACCCAGAGCCCCAGCAGCCTGCCTGCCTCTCTGGGCGATAGAGTGACCATCAACTGCCAGGCCAGCCAGGACATCAGCAACTACCTGAACTGGTATCAGCAGAAGCCCGGCAAGGCCCCCAAGCTGCTGATCTACTACACCAACAAGCTGGCCGACGGTGTGCCCAGCAGATTCAGCGGCAGCGGTAGCGGCAGAGACAGCAGCTTCACCATCAGCTCCCTGGAATCCGAGGATATCGGCAGCTACTACTGCCAGCAGTACTACAACTACCCCTGGACCTTCGGCCCTGGCACCAAGCTGGAAATCAAAAGAGGAGGTGGTGGATCCCAGGTGCAGCTGAAACAGAGCGGCCCGGGCCTGGTGCAGCCGAGCCAGAGCCTGAGCATTACCTGCACCGTGAGCGGCTTTAGCCTGACCAACTATGGCGTGCATTGGGTGCGCCAGAGCCCGGGCAAAGGCCTGGAATGGCTGGGCGTGATTTGGAGCGGCGGCAACACCGATTATAACACCCCGTTTACCAGCCGCCTGAGCATTAACAAAGATAACAGCAAAAGCCAGGTGTTTTTTAAAATGAACAGCCTGCAAAGCCAGGATACCGCGATTTATTATTGCGCGCGCGCGCTGACCTATTATGATTATGAATTTGCGTATTGGGGCCAGGGCACCCTGGTGACCGTGAGCGCGGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA] Amino Acid Sequence[spacer (SEQ ID NO: 87)][pLW121 without spacer (SEQ ID NO: 540)](SEQ ID NO: 486)[QGQSGQ][IHPMCHPEGVCVALDGGGSSGGSGGSGGISSGLLSGRSDNHGGGSEVQLVESGGGLVQPGKSLKLSCEASGFTFSGYGMHWVRQAPGRGLESVAYITSSSINIKYADAVKGRFTVSRDNAKNLLFLQMNILKSEDTAMYYCARFDWDKNYWGQGTMVTVSSGGGGSGGGSGGSGGGSGGGGSDIQMTQSPSSLPASLGDRVTINCQASQDISNYLNWYQQKPGKAPKLLIYYTNKLADGVPSRFSGSGSGRDSSFTISSLESEDIGSYYCQQYYNYPWTFGPGTKLEIKRGGGGSQVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSQDTAIYYCARALTYYDYEFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]*OPP022: LC C225 3954-1204 Nucleotide Sequence[spacer (SEQ ID NO: 507)][OPP022 without spacer (SEQ ID NO: 509)](SEQ ID NO: 449)[CAAGGCCAGTCTGGCCAG][TGCATCTCACCTCGTGGTTGTCCGGACGGCCCATACGTCATGTACGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGATCCGGTCTGAGCGGCCGTTCCGATAATCATGGCAGTAGCGGTACCCAGATCTTGCTGACCCAGAGCCCGGTGATTCTGAGCGTGAGCCCGGGCGAACGTGTGAGCTTTAGCTGCCGCGCGAGCCAGAGCATTGGCACCAACATTCATTGGTATCAGCAGCGCACCAACGGCAGCCCGCGCCTGCTGATTAAATATGCGAGCGAAAGCATTAGCGGCATTCCGAGCCGCTTTAGCGGCAGCGGCAGCGGCACCGATTTTACCCTGAGCATTAACAGCGTGGAAAGCGAAGATATTGCGGATTATTATTGCCAGCAGAACAACAACTGGCCGACCACCTTTGGCGCGGGCACCAAACTGGAACTGAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG] Amino Acid Sequence[spacer (SEQ ID NO: 87)][OPP022 without spacer (SEQ ID NO: 508)](SEQ ID NO: 450)[QGQSGQ][CISPRGCPDGPYVMYGSSGGSGGSGGSGLSGRSDNHGSSGTQILLTQSPVILSVSPGERVSFSCRASQSIGTNIHWYQQRTNGSPRLLIKYASESISGIPSRFSGSGSGTDFTLSINSVESEDIADYYCQQNNNWPTTFGAGTKLELKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC]*CI050: 3954-1204-C225v5N297Q-MC06-2001-mCD3-H-NpLW122: HC C225v5N297Q-MC06-2001-mCD3-H-N Nucleotide Sequence[spacer (SEQ ID NO: 507)][pLW122 without spacer (SEQ ID NO: 541)](SEQ ID NO: 487)[CAAGGCCAGTCTGGCCAA][GATTGCTTTGTGCCTGGGTGGTATTTGGCGGGTCCGTGCGCTCAGGGAGGTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTATTAGCAGTGGTCTGTTAAGCGGTCGTAGCGATAATCATGGCGGCGGTTCTGAAGTGCAGCTGGTGGAATCTGGGGGCGGACTGGTGCAGCCTGGCAAGTCTCTGAAGCTGAGCTGCGAGGCCAGCGGCTTCACCTTTAGCGGCTACGGCATGCACTGGGTGCGCCAGGCACCTGGCAGAGGCCTGGAAAGCGTGGCCTACATCACCAGCAGCAGCATCAACATTAAGTACGCCGACGCCGTGAAGGGCCGGTTCACCGTGTCCAGAGACAACGCCAAGAACCTGCTGTTCCTGCAGATGAACATCCTGAAGTCCGAGGACACCGCCATGTACTACTGCGCCAGATTCGACTGGGACAAGAACTACTGGGGCCAGGGCACAATGGTCACAGTGTCCAGCGGTGGAGGTGGTAGTGGTGGAGGAAGTGGAGGTTCAGGAGGTGGAAGCGGTGGTGGTGGTAGTGACATCCAGATGACCCAGAGCCCCAGCAGCCTGCCTGCCTCTCTGGGCGATAGAGTGACCATCAACTGCCAGGCCAGCCAGGACATCAGCAACTACCTGAACTGGTATCAGCAGAAGCCCGGCAAGGCCCCCAAGCTGCTGATCTACTACACCAACAAGCTGGCCGACGGTGTGCCCAGCAGATTCAGCGGCAGCGGTAGCGGCAGAGACAGCAGCTTCACCATCAGCTCCCTGGAATCCGAGGATATCGGCAGCTACTACTGCCAGCAGTACTACAACTACCCCTGGACCTTCGGCCCTGGCACCAAGCTGGAAATCAAAAGAGGAGGTGGTGGATCCCAGGTGCAGCTGAAACAGAGCGGCCCGGGCCTGGTGCAGCCGAGCCAGAGCCTGAGCATTACCTGCACCGTGAGCGGCTTTAGCCTGACCAACTATGGCGTGCATTGGGTGCGCCAGAGCCCGGGCAAAGGCCTGGAATGGCTGGGCGTGATTTGGAGCGGCGGCAACACCGATTATAACACCCCGTTTACCAGCCGCCTGAGCATTAACAAAGATAACAGCAAAAGCCAGGTGTTTTTTAAAATGAACAGCCTGCAAAGCCAGGATACCGCGATTTATTATTGCGCGCGCGCGCTGACCTATTATGATTATGAATTTGCGTATTGGGGCCAGGGCACCCTGGTGACCGTGAGCGCGGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA] Amino Acid Sequence[spacer (SEQ ID NO: 87)][pLW122 without spacer (SEQ ID NO: 542)](SEQ ID NO: 488)[QGQSGQ][DCFVPGWYLAGPCAQGGGSSGGSGGSGGISSGLLSGRSDNHGGGSEVQLVESGGGLVQPGKSLKLSCEASGFTFSGYGMHWVRQAPGRGLESVAYITSSSINIKYADAVKGRFTVSRDNAKNLLFLQMNILKSEDTAMYYCARFDWDKNYWGQGTMVTVSSGGGGSGGGSGGSGGGSGGGGSDIQMTQSPSSLPASLGDRVTINCQASQDISNYLNWYQQKPGKAPKLLIYYTNKLADGVPSRFSGSGSGRDSSFTISSLESEDIGSYYCQQYYNYPWTFGPGTKLEIKRGGGGSQVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSQDTAIYYCARALTYYDYEFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]* OPP022: LC C225 3954-1204 Nucleotide Sequence[spacer (SEQ ID NO: 507)][OPP022 without spacer (SEQ ID NO: 509)](SEQ ID NO: 449)[CAAGGCCAGTCTGGCCAG][TGCATCTCACCTCGTGGTTGTCCGGACGGCCCATACGTCATGTACGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGATCCGGTCTGAGCGGCCGTTCCGATAATCATGGCAGTAGCGGTACCCAGATCTTGCTGACCCAGAGCCCGGTGATTCTGAGCGTGAGCCCGGGCGAACGTGTGAGCTTTAGCTGCCGCGCGAGCCAGAGCATTGGCACCAACATTCATTGGTATCAGCAGCGCACCAACGGCAGCCCGCGCCTGCTGATTAAATATGCGAGCGAAAGCATTAGCGGCATTCCGAGCCGCTTTAGCGGCAGCGGCAGCGGCACCGATTTTACCCTGAGCATTAACAGCGTGGAAAGCGAAGATATTGCGGATTATTATTGCCAGCAGAACAACAACTGGCCGACCACCTTTGGCGCGGGCACCAAACTGGAACTGAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG] Amino Acid Sequence[spacer (SEQ ID NO: 87)][OPP022 without spacer (SEQ ID NO: 508)](SEQ ID NO: 450)[QGQSGQ][CISPRGCPDGPYVMYGSSGGSGGSGGSGLSGRSDNHGSSGTQILLTQSPVILSVSPGERVSFSCRASQSIGTNIHWYQQRTNGSPRLLIKYASESISGIPSRFSGSGSGTDFTLSINSVESEDIADYYCQQNNNWPTTFGAGTKLELKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC]*CI051: 3954-1204-C225v5N297Q-MC07-2001-mCD3-H-NpLW123: HC C225v5N297Q-MC07-2001-mCD3-H-N Nucleotide Sequence[spacer (SEQ ID NO: 521)][pLW123 without spacer (SEQ ID NO: 543)](SEQ ID NO: 489)[CAAGGCCAGTCTGGCCAAGGT][GTGTGCCATTCTCGGTTGGAGTGGCTTCTGGGTTGCCAAGGAGGAGGTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTATTAGCAGTGGTCTGTTAAGCGGTCGTAGCGATAATCATGGCGGCGGTTCTGAAGTGCAGCTGGTGGAATCTGGGGGCGGACTGGTGCAGCCTGGCAAGTCTCTGAAGCTGAGCTGCGAGGCCAGCGGCTTCACCTTTAGCGGCTACGGCATGCACTGGGTGCGCCAGGCACCTGGCAGAGGCCTGGAAAGCGTGGCCTACATCACCAGCAGCAGCATCAACATTAAGTACGCCGACGCCGTGAAGGGCCGGTTCACCGTGTCCAGAGACAACGCCAAGAACCTGCTGTTCCTGCAGATGAACATCCTGAAGTCCGAGGACACCGCCATGTACTACTGCGCCAGATTCGACTGGGACAAGAACTACTGGGGCCAGGGCACAATGGTCACAGTGTCCAGCGGTGGAGGTGGTAGTGGTGGAGGAAGTGGAGGTTCAGGAGGTGGAAGCGGTGGTGGTGGTAGTGACATCCAGATGACCCAGAGCCCCAGCAGCCTGCCTGCCTCTCTGGGCGATAGAGTGACCATCAACTGCCAGGCCAGCCAGGACATCAGCAACTACCTGAACTGGTATCAGCAGAAGCCCGGCAAGGCCCCCAAGCTGCTGATCTACTACACCAACAAGCTGGCCGACGGTGTGCCCAGCAGATTCAGCGGCAGCGGTAGCGGCAGAGACAGCAGCTTCACCATCAGCTCCCTGGAATCCGAGGATATCGGCAGCTACTACTGCCAGCAGTACTACAACTACCCCTGGACCTTCGGCCCTGGCACCAAGCTGGAAATCAAAAGAGGAGGTGGTGGATCCCAGGTGCAGCTGAAACAGAGCGGCCCGGGCCTGGTGCAGCCGAGCCAGAGCCTGAGCATTACCTGCACCGTGAGCGGCTTTAGCCTGACCAACTATGGCGTGCATTGGGTGCGCCAGAGCCCGGGCAAAGGCCTGGAATGGCTGGGCGTGATTTGGAGCGGCGGCAACACCGATTATAACACCCCGTTTACCAGCCGCCTGAGCATTAACAAAGATAACAGCAAAAGCCAGGTGTTTTTTAAAATGAACAGCCTGCAAAGCCAGGATACCGCGATTTATTATTGCGCGCGCGCGCTGACCTATTATGATTATGAATTTGCGTATTGGGGCCAGGGCACCCTGGTGACCGTGAGCGCGGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA] Amino Acid Sequence[spacer (SEQ ID NO: 407)][pLW123 without spacer (SEQ ID NO: 544)](SEQ ID NO: 490)[QGQSGQG][VCHSRLEWLLGCQGGGGSSGGSGGSGGISSGLLSGRSDNHGGGSEVQLVESGGGLVQPGKSLKLSCEASGFTFSGYGMHWVRQAPGRGLESVAYITSSSINIKYADAVKGRFTVSRDNAKNLLFLQMNILKSEDTAMYYCARFDWDKNYWGQGTMVTVSSGGGGSGGGSGGSGGGSGGGGSDIQMTQSPSSLPASLGDRVTINCQASQDISNYLNWYQQKPGKAPKLLIYYTNKLADGVPSRFSGSGSGRDSSFTISSLESEDIGSYYCQQYYNYPWTFGPGTKLEIKRGGGGSQVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSQDTAIYYCARALTYYDYEFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]*OPP022: LC C225 3954-1204 Nucleotide Sequence[spacer (SEQ ID NO: 507)][OPP022 without spacer (SEQ ID NO: 509)](SEQ ID NO: 449)[CAAGGCCAGTCTGGCCAG][TGCATCTCACCTCGTGGTTGTCCGGACGGCCCATACGTCATGTACGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGATCCGGTCTGAGCGGCCGTTCCGATAATCATGGCAGTAGCGGTACCCAGATCTTGCTGACCCAGAGCCCGGTGATTCTGAGCGTGAGCCCGGGCGAACGTGTGAGCTTTAGCTGCCGCGCGAGCCAGAGCATTGGCACCAACATTCATTGGTATCAGCAGCGCACCAACGGCAGCCCGCGCCTGCTGATTAAATATGCGAGCGAAAGCATTAGCGGCATTCCGAGCCGCTTTAGCGGCAGCGGCAGCGGCACCGATTTTACCCTGAGCATTAACAGCGTGGAAAGCGAAGATATTGCGGATTATTATTGCCAGCAGAACAACAACTGGCCGACCACCTTTGGCGCGGGCACCAAACTGGAACTGAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG] Amino Acid Sequence[spacer (SEQ ID NO: 87)][OPP022 without spacer (SEQ ID NO: 508)](SEQ ID NO: 450)[QGQSGQ][CISPRGCPDGPYVMYGSSGGSGGSGGSGLSGRSDNHGSSGTQILLTQSPVILSVSPGERVSFSCRASQSIGTNIHWYQQRTNGSPRLLIKYASESISGIPSRFSGSGSGTDFTLSINSVESEDIADYYCQQNNNWPTTFGAGTKLELKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC]*CI057: 3954-1204-C225v5N297Q-MC08-2001-mCD3-H-NpLW124: HC C225v5N297Q-MC08-2001-mCD3-H-N Nucleotide Sequence[spacer (SEQ ID NO: 507)][pLW124 without spacer (SEQ ID NO: 545)](SEQ ID NO: 501)[CAAGGCCAGTCTGGCCAA][GTGGGTGAGTGCGTTCCGGGTCCGCATGGGTGCTGGATGGCTTATGGAGGTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTATTAGCAGTGGTCTGTTAAGCGGTCGTAGCGATAATCATGGCGGCGGTTCTGAAGTGCAGCTGGTGGAATCTGGGGGCGGACTGGTGCAGCCTGGCAAGTCTCTGAAGCTGAGCTGCGAGGCCAGCGGCTTCACCTTTAGCGGCTACGGCATGCACTGGGTGCGCCAGGCACCTGGCAGAGGCCTGGAAAGCGTGGCCTACATCACCAGCAGCAGCATCAACATTAAGTACGCCGACGCCGTGAAGGGCCGGTTCACCGTGTCCAGAGACAACGCCAAGAACCTGCTGTTCCTGCAGATGAACATCCTGAAGTCCGAGGACACCGCCATGTACTACTGCGCCAGATTCGACTGGGACAAGAACTACTGGGGCCAGGGCACAATGGTCACAGTGTCCAGCGGTGGAGGTGGTAGTGGTGGAGGAAGTGGAGGTTCAGGAGGTGGAAGCGGTGGTGGTGGTAGTGACATCCAGATGACCCAGAGCCCCAGCAGCCTGCCTGCCTCTCTGGGCGATAGAGTGACCATCAACTGCCAGGCCAGCCAGGACATCAGCAACTACCTGAACTGGTATCAGCAGAAGCCCGGCAAGGCCCCCAAGCTGCTGATCTACTACACCAACAAGCTGGCCGACGGTGTGCCCAGCAGATTCAGCGGCAGCGGTAGCGGCAGAGACAGCAGCTTCACCATCAGCTCCCTGGAATCCGAGGATATCGGCAGCTACTACTGCCAGCAGTACTACAACTACCCCTGGACCTTCGGCCCTGGCACCAAGCTGGAAATCAAAAGAGGAGGTGGTGGATCCCAGGTGCAGCTGAAACAGAGCGGCCCGGGCCTGGTGCAGCCGAGCCAGAGCCTGAGCATTACCTGCACCGTGAGCGGCTTTAGCCTGACCAACTATGGCGTGCATTGGGTGCGCCAGAGCCCGGGCAAAGGCCTGGAATGGCTGGGCGTGATTTGGAGCGGCGGCAACACCGATTATAACACCCCGTTTACCAGCCGCCTGAGCATTAACAAAGATAACAGCAAAAGCCAGGTGTTTTTTAAAATGAACAGCCTGCAAAGCCAGGATACCGCGATTTATTATTGCGCGCGCGCGCTGACCTATTATGATTATGAATTTGCGTATTGGGGCCAGGGCACCCTGGTGACCGTGAGCGCGGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA] Amino Acid Sequence[spacer (SEQ ID NO: 87)][pLW124 without spacer (SEQ ID NO: 546)](SEQ ID NO: 502)[QGQSGQ][VGECVPGPHGCWMAYGGGSSGGSGGSGGISSGLLSGRSDNHGGGSEVQLVESGGGLVQPGKSLKLSCEASGFTFSGYGMHWVRQAPGRGLESVAYITSSSINIKYADAVKGRFTVSRDNAKNLLFLQMNILKSEDTAMYYCARFDWDKNYWGQGTMVTVSSGGGGSGGGSGGSGGGSGGGGSDIQMTQSPSSLPASLGDRVTINCQASQDISNYLNWYQQKPGKAPKLLIYYTNKLADGVPSRFSGSGSGRDSSFTISSLESEDIGSYYCQQYYNYPWTFGPGTKLEIKRGGGGSQVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSQDTAIYYCARALTYYDYEFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]*OPP022: LC C225 3954-1204 Nucleotide Sequence[spacer (SEQ ID NO: 507)][OPP022 without spacer (SEQ ID NO: 509)](SEQ ID NO: 449)[CAAGGCCAGTCTGGCCAG][TGCATCTCACCTCGTGGTTGTCCGGACGGCCCATACGTCATGTACGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGATCCGGTCTGAGCGGCCGTTCCGATAATCATGGCAGTAGCGGTACCCAGATCTTGCTGACCCAGAGCCCGGTGATTCTGAGCGTGAGCCCGGGCGAACGTGTGAGCTTTAGCTGCCGCGCGAGCCAGAGCATTGGCACCAACATTCATTGGTATCAGCAGCGCACCAACGGCAGCCCGCGCCTGCTGATTAAATATGCGAGCGAAAGCATTAGCGGCATTCCGAGCCGCTTTAGCGGCAGCGGCAGCGGCACCGATTTTACCCTGAGCATTAACAGCGTGGAAAGCGAAGATATTGCGGATTATTATTGCCAGCAGAACAACAACTGGCCGACCACCTTTGGCGCGGGCACCAAACTGGAACTGAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG] Amino Acid Sequence[spacer (SEQ ID NO: 87)][OPP022 without spacer (SEQ ID NO: 508)](SEQ ID NO: 450)[QGQSGQ][CISPRGCPDGPYVMYGSSGGSGGSGGSGLSGRSDNHGSSGTQILLTQSPVILSVSPGERVSFSCRASQSIGTNIHWYQQRTNGSPRLLIKYASESISGIPSRFSGSGSGTDFTLSINSVESEDIADYYCQQNNNWPTTFGAGTKLELKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC]*CI058: 3954-1204-C225v5N297Q-MC09-2001-mCD3-H-NpLW125: HC C225v5N297Q-MC09-2001-mCD3-H-N Nucleotide Sequence[spacer (SEQ ID NO: 507)][pLW125 without spacer (SEQ ID NO: 475)](SEQ ID NO: 503)[CAAGGCCAGTCTGGCCAA][TGGACGTGCGGAGGTATGGTGTATCTTGCGGGATTCTGCATGGCGGGAGGTGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGTATTAGCAGTGGTCTGTTAAGCGGTCGTAGCGATAATCATGGCGGCGGTTCTGAAGTGCAGCTGGTGGAATCTGGGGGCGGACTGGTGCAGCCTGGCAAGTCTCTGAAGCTGAGCTGCGAGGCCAGCGGCTTCACCTTTAGCGGCTACGGCATGCACTGGGTGCGCCAGGCACCTGGCAGAGGCCTGGAAAGCGTGGCCTACATCACCAGCAGCAGCATCAACATTAAGTACGCCGACGCCGTGAAGGGCCGGTTCACCGTGTCCAGAGACAACGCCAAGAACCTGCTGTTCCTGCAGATGAACATCCTGAAGTCCGAGGACACCGCCATGTACTACTGCGCCAGATTCGACTGGGACAAGAACTACTGGGGCCAGGGCACAATGGTCACAGTGTCCAGCGGTGGAGGTGGTAGTGGTGGAGGAAGTGGAGGTTCAGGAGGTGGAAGCGGTGGTGGTGGTAGTGACATCCAGATGACCCAGAGCCCCAGCAGCCTGCCTGCCTCTCTGGGCGATAGAGTGACCATCAACTGCCAGGCCAGCCAGGACATCAGCAACTACCTGAACTGGTATCAGCAGAAGCCCGGCAAGGCCCCCAAGCTGCTGATCTACTACACCAACAAGCTGGCCGACGGTGTGCCCAGCAGATTCAGCGGCAGCGGTAGCGGCAGAGACAGCAGCTTCACCATCAGCTCCCTGGAATCCGAGGATATCGGCAGCTACTACTGCCAGCAGTACTACAACTACCCCTGGACCTTCGGCCCTGGCACCAAGCTGGAAATCAAAAGAGGAGGTGGTGGATCCCAGGTGCAGCTGAAACAGAGCGGCCCGGGCCTGGTGCAGCCGAGCCAGAGCCTGAGCATTACCTGCACCGTGAGCGGCTTTAGCCTGACCAACTATGGCGTGCATTGGGTGCGCCAGAGCCCGGGCAAAGGCCTGGAATGGCTGGGCGTGATTTGGAGCGGCGGCAACACCGATTATAACACCCCGTTTACCAGCCGCCTGAGCATTAACAAAGATAACAGCAAAAGCCAGGTGTTTTTTAAAATGAACAGCCTGCAAAGCCAGGATACCGCGATTTATTATTGCGCGCGCGCGCTGACCTATTATGATTATGAATTTGCGTATTGGGGCCAGGGCACCCTGGTGACCGTGAGCGCGGCTAGCACCAAGGGCCCATCGGTCTTCCCCCTGGCACCCTCCTCCAAGAGCACCTCTGGGGGCACAGCGGCCCTGGGCTGCCTGGTCAAGGACTACTTCCCCGAACCGGTGACGGTGTCGTGGAACTCAGGCGCCCTGACCAGCGGCGTGCACACCTTCCCGGCTGTCCTACAGTCCTCAGGACTCTACTCCCTCAGCAGCGTGGTGACCGTGCCCTCCAGCAGCTTGGGCACCCAGACCTACATCTGCAACGTGAATCACAAGCCCAGCAACACCAAGGTGGACAAGAAAGTTGAGCCCAAATCTTGTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAACTCCTGGGGGGACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACCCCTGAGGTCACATGCGTGGTGGTGGACGTGAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACCAGAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAGCCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGAGGAGATGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCCAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGA] Amino Acid Sequence[spacer (SEQ ID NO: 87)][pLW087 without spacer (SEQ ID NO: 476)](SEQ ID NO: 504)[QGQSGQ][WTCGGMVYLAGFCMAGGGSSGGSGGSGGISSGLLSGRSDNHGGGSEVQLVESGGGLVQPGKSLKLSCEASGFTFSGYGMHWVRQAPGRGLESVAYITSSSINIKYADAVKGRFTVSRDNAKNLLFLQMNILKSEDTAMYYCARFDWDKNYWGQGTMVTVSSGGGGSGGGSGGSGGGSGGGGSDIQMTQSPSSLPASLGDRVTINCQASQDISNYLNWYQQKPGKAPKLLIYYTNKLADGVPSRFSGSGSGRDSSFTISSLESEDIGSYYCQQYYNYPWTFGPGTKLEIKRGGGGSQVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSQDTAIYYCARALTYYDYEFAYWGQGTLVTVSAASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK]*OPP022: LC C225 3954-1204 Nucleotide Sequence[spacer (SEQ ID NO: 507)][OPP022 without spacer (SEQ ID NO: 509)](SEQ ID NO: 449)[CAAGGCCAGTCTGGCCAG][TGCATCTCACCTCGTGGTTGTCCGGACGGCCCATACGTCATGTACGGCTCGAGCGGTGGCAGCGGTGGCTCTGGTGGATCCGGTCTGAGCGGCCGTTCCGATAATCATGGCAGTAGCGGTACCCAGATCTTGCTGACCCAGAGCCCGGTGATTCTGAGCGTGAGCCCGGGCGAACGTGTGAGCTTTAGCTGCCGCGCGAGCCAGAGCATTGGCACCAACATTCATTGGTATCAGCAGCGCACCAACGGCAGCCCGCGCCTGCTGATTAAATATGCGAGCGAAAGCATTAGCGGCATTCCGAGCCGCTTTAGCGGCAGCGGCAGCGGCACCGATTTTACCCTGAGCATTAACAGCGTGGAAAGCGAAGATATTGCGGATTATTATTGCCAGCAGAACAACAACTGGCCGACCACCTTTGGCGCGGGCACCAAACTGGAACTGAAACGTACGGTGGCTGCACCATCTGTCTTCATCTTCCCGCCATCTGATGAGCAGTTGAAATCTGGAACTGCCTCTGTTGTGTGCCTGCTGAATAACTTCTATCCCAGAGAGGCCAAAGTACAGTGGAAGGTGGATAACGCCCTCCAATCGGGTAACTCCCAGGAGAGTGTCACAGAGCAGGACAGCAAGGACAGCACCTACAGCCTCAGCAGCACCCTGACGCTGAGCAAAGCAGACTACGAGAAACACAAAGTCTACGCCTGCGAAGTCACCCATCAGGGCCTGAGCTCGCCCGTCACAAAGAGCTTCAACAGGGGAGAGTGTTAG] Amino Acid Sequence[spacer (SEQ ID NO: 87)][OPP022 without spacer (SEQ ID NO: 508)](SEQ ID NO: 450)[QGQSGQ][CISPRGCPDGPYVMYGSSGGSGGSGGSGLSGRSDNHGSSGTQILLTQSPVILSVSPGERVSFSCRASQSIGTNIHWYQQRTNGSPRLLIKYASESISGIPSRFSGSGSGTDFTLSINSVESEDIADYYCQQNNNWPTTFGAGTKLELKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC]* CI059: BiTE420 BiTE420Nucleotide Sequence (SEQ ID NO: 505)GACATCTTGCTGACTCAGTCTCCAGTCATCCTGTCTGTGAGTCCAGGAGAAAGAGTCAGTTTCTCCTGCAGGGCCAGTCAGAGCATTGGCACAAACATACACTGGTATCAGCAAAGAACAAATGGTTCTCCAAGGCTTCTCATAAAGTATGCTTCTGAGTCTATCTCTGGGATCCCTTCCAGGTTTAGTGGCAGTGGATCAGGGACAGATTTTACTCTTAGCATCAACAGTGTGGAGTCTGAAGATATTGCAGATTATTACTGTCAACAAAATAATAACTGGCCAACCACATTTGGTGCAGGAACAAAGCTGGAACTGAAAGGTGGTGGTGGTTCTGGCGGCGGCGGCTCCGGTGGTGGTGGTTCTCAGGTGCAGCTGAAGCAGTCAGGACCTGGCCTAGTGCAGCCCTCACAGAGCCTGTCCATCACCTGCACAGTCTCTGGTTTCTCATTAACTAACTATGGAGTACACTGGGTTCGCCAGTCTCCAGGAAAGGGTCTGGAGTGGCTGGGAGTGATATGGAGTGGTGGAAACACAGACTATAATACACCTTTCACATCCAGACTGAGCATCAACAAGGACAATTCCAAGAGCCAAGTTTTCTTTAAAATGAACAGTCTGCAATCTAATGACACAGCCATATATTACTGTGCCAGAGCCCTGACCTATTATGACTACGAGTTCGCCTATTGGGGTCAGGGAACCCTGGTTACCGTGTCTTCCGGAGGTGGTGGATCCGAGGTGCAGCTGGTCGAGTCTGGAGGAGGATTGGTGCAGCCTGGAGGGTCATTGAAACTCTCATGTGCAGCCTCTGGATTCACCTTCAATAAGTACGCCATGAACTGGGTCCGCCAGGCTCCAGGAAAGGGTTTGGAATGGGTTGCTCGCATAAGAAGTAAATATAATAATTATGCAACATATTATGCCGATTCAGTGAAAGACAGGTTCACCATCTCCAGAGATGATTCAAAAAACACTGCCTATCTACAAATGAACAACTTGAAAACTGAGGACACTGCCGTGTACTACTGTGTGAGACATGGGAACTTCGGTAATAGCTACATATCCTACTGGGCTTACTGGGGCCAAGGGACTCTGGTCACCGTCTCCTCAGGTGGTGGTGGTTCTGGCGGCGGCGGCTCCGGTGGTGGTGGTTCTCAGACTGTTGTGACTCAGGAACCTTCACTCACCGTATCACCTGGTGGAACAGTCACACTCACTTGTGGCTCCTCGACTGGGGCTGTTACATCTGGCTACTACCCAAACTGGGTCCAACAAAAACCAGGTCAGGCACCCCGTGGTCTAATAGGTGGGACTAAGTTCCTCGCCCCCGGTACTCCTGCCAGATTCTCAGGCTCCCTGCTTGGAGGCAAGGCTGCCCTCACCCTCTCAGGGGTACAGCCAGAGGATGAGGCAGAATATTACTGTGCTCTATGGTACAGCAACCGCTGGGTGTTCGGTGGAGGAACCAAACTGACTGTCCTACACCATCACCACCATCATCACCACTAG  Amino Acid Sequence(SEQ ID NO: 506)DILLTQSPVILSVSPGERVSFSCRASQSIGTNIHWYQQRTNGSPRLLIKYASESISGIPSRFSGSGSGTDFTLSINSVESEDIADYYCQQNNNWPTTFGAGTKLELKGGGGSGGGGSGGGGSQVQLKQSGPGLVQPSQSLSITCTVSGFSLTNYGVHWVRQSPGKGLEWLGVIWSGGNTDYNTPFTSRLSINKDNSKSQVFFKMNSLQSNDTAIYYCARALTYYDYEFAYWGQGTLVTVSSGGGGSEVQLVESGGGLVQPGGSLKLSCAASGFTFNKYAMNWVRQAPGKGLEWVARIRSKYNNYATYYADSVKDRFTISRDDSKNTAYLQMNNLKTEDTAVYYCVRHGNFGNSYISYWAYWGQGTLVTVSSGGGGSGGGGSGGGGSQTVVTQEPSLTVSPGGTVTLTCGSSTGAVTSGYYPNWVQQKPGQAPRGLIGGTKFLAPGTPARFSGSLLGGKAALTLSGVQPEDEAEYYCALWYSNRWVFGGGTKLTVL* 

Example 5 SP34 scFv-Fc Binding to CD3ε Positive Jurkat Cells

To determine if the SP34 scFv Fc fusions could bind to CD3ε positiveJurkat T cells, a flow cytometry-based binding assay was performed.Jurkat T cells (Clone E6-1, ATCC, TIB-152) were cultured inRPMI-1640+glutamax (Life Technologies, Catalog 72400-120), 10% HeatInactivated-Fetal Bovine Serum (HI-FBS, Life Technologies, Catalog10438-026), 100 U/ml penicillin, and 100 μg/ml streptomycin (LifeTechnologies, Catalog 15140-122) according to ATCC guidelines. Cellswere harvested by centrifugation (200×g, 4° C., 5 min) and re-suspendedin PBS supplemented with 2% HI-FBS (FACS Buffer). About 250,000 Jurkatsper well were transferred to a 96-well U-bottom plate, harvested, andre-suspended in 50 μL of primary antibody. The starting concentration ofantibody was 200 nM followed by 5-fold serial dilutions for a total of 8concentrations. The following antibodies were tested: SP34 LvHv scFv Fc,SP34 HvLv scFv Fc, and SP34-2.

Cells were incubated at 4° C. with shaking for about 2 hours, harvested,and washed with 3×200 μL of FACS Buffer. Jurkats treated with the SP34scFv-Fc fusions were resuspended in 50 μl Alexa Fluor 488 conjugatedanti-Human IgG Fc (1:100 dilution, Jackson ImmunoResearch, Product109-546-098). Cells treated with SP34-2 and mouse isotype wereresuspended in 50 μl Alexa Fluor 647 conjugated Anti-mouse IgG (1:100dilution, Jackson ImmunoResearch, Product 715-605-150). Jurkats wereincubated at 4° C. with shaking for about 30 min, harvested, washed with3×200 μL of FACS Buffer, and resuspended in a final volume of 120 μL ofFACS Buffer. Samples were analyzed on a BD Accuri C6 (BD Biosciences)and the median fluorescence intensity (MFI) of viable cells wascalculated using FlowJo V10 (Treestar). EC₅₀ values were calculated inGraphPad Prism 6 by curve fitting the data to log(agonist) vs. response(three parameters).

FIGS. 13A and 13B demonstrate that all the formats of SP34 bind to CD3εJurkat cells with similar EC₅₀ values.

Example 6 Binding of Activatable Anti-CD3ε Antibodies to CD3ε JurkatCells

To determine if the masking peptides described in the previous examplescould inhibit binding in the SP34 scFv-Fc context, a flowcytometry-based binding assay was performed. Jurkat T cells werecultured and processed in FACS Buffer as described. About 100,000Jurkats per well were transferred to a 96-well U-bottom plate,harvested, and re-suspended in 50 μL of primary antibody. Titrationsstarted at either 1 μM or 333 nM followed by 3-fold serial dilutions inFACS Buffer. The following antibodies and activatable antibodies weretested: 15003 1204 SP34 LvHv Fc; 15860 1204 SP34 LvHv Fc; 15865 1204SP34 LvHv, SP34 LvHv, 15003 1204 SP34 HvLv; 15860 1204 SP34 HvLv Fc;15865 1204 SP34 HvLv, SP34 HvLv Fc, and mouse SP34.

Cells were incubated at 4° C. with shaking for about 1 hour, harvested,and washed with 3×200 μL of FACS Buffer. Jurkats treated with SP34 Fcfusions were resuspended in 50 μl of Alexa Fluor 647 conjugatedanti-Human IgG Fc (1:100 dilution, Jackson ImmunoResearch, Product109-606-008). Cells treated with mouse SP34 were resuspended in 50 μlAlexa Fluor 647 conjugated Anti-mouse IgG (1:100 dilution, JacksonImmunoResearch, Product 715-605-150). Jurkats were incubated at 4° C.with shaking for about 30 min, harvested, washed with 3×200 μL of FACSBuffer, and resuspended in a final volume of 100 μL of FACS Buffercontaining 2.5 μg/ml 7-AAD (BD Biosciences, Catalog 559925). Sampleswere collected on a MACSQuant® Analyzer 10 (Miltenyi) and the medianfluorescence intensity (MFI) of viable cells was calculated using FlowJoV10 (Treestar). EC₅₀ values were calculated in GraphPad Prism 6 by curvefitting the data to log(agonist) vs. response (three parameters).

FIGS. 14A and 14B demonstrate that incorporation of the masking peptidesinto the SP34 scFv-Fc context to produce activatable antibodies shiftedthe EC₅₀ values for CD3ε binding from single digit to triple digit nM.

Example 7 Binding of Dually Masked Multispecific, Activatable Antibodiesto EGFR+ HT-29 Cells and CD3ε+ Jurkat Cells

To determine if the CD3ε masking peptides described in the previousexamples could inhibit binding in the context of an EGFR maskedmultispecific, activatable antibody, a flow cytometry-based bindingassay was performed.

EGFR+ HT-29-luc2 (Caliper) were cultured in RPMI-1640+glutamax (LifeTechnologies, Catalog 72400-120), 10% Heat Inactivated-Fetal BovineSerum (HI-FBS, Life Technologies, Catalog 10438-026), 100 U/mlpenicillin, and 100 μg/ml streptomycin (Life Technologies, Catalog15140-122) according to manufacturer guidelines. Cells were lifted withcell dissociation buffer (Sigma, Catalog C5789), washed, andre-suspended in 50 μL of primary antibody. Titrations started at theconcentrations indicated in FIGS. 15A-15D followed by 3-fold serialdilutions in FACS Buffer. The following multispecific antibodies andmultispecific, activatable antibodies were tested: CI005 (EGFR/hCD3),CI023 (M-EGFR/M-hCD3), CI024 (M-EGFR/M-hCD3), CI015 (EGFR/hCD3), CI011(M-EGFR/M-hCD3), CI010 (M-EGFR/M-hCD3). Two versions of SP34 scFv areutilized, namely the scFv in CI005, CI023, CI024 versus the scFv inCI015, CI011, CI010. The binding specificity of the multispecificantibody is indicated by antigen name (in this example EGFR and CD3) andthe letter M indicates masking of antigen binding.

HT29-luc2 cells were incubated at 4° C. with shaking for about 1 hour,harvested, and washed with 3×200 μL of FACS Buffer. Cells wereresuspended in 50 μl of Alexa Fluor 647 conjugated anti-Human IgG Fc(1:100 dilution, Jackson ImmunoResearch, Product 109-606-008) andincubated at 4° C. with shaking for about 30 min. HT29-luc2 wereharvested, washed with 3×200 μL of FACS Buffer, and resuspended in afinal volume of 100 μL of FACS Buffer containing 2.5 μg/ml 7-AAD (BDBiosciences, Catalog 559925). Samples were collected on a MACSQuant®Analyzer 10 (Miltenyi) and the median fluorescence intensity (MFI) ofviable cells was calculated using FlowJo V10 (Treestar). EC₅₀ valueswere calculated in GraphPad Prism 6 by curve fitting the data tolog(agonist) vs. response (three parameters).

Binding of the multispecific antibodies and multispecific, activatableantibodies to CD3ε Jurkats were tested as previously described.

FIGS. 15A-15C demonstrate that incorporation of the CD3ε maskingpeptides into the EGFR masked multispecific activatable antibody shiftedthe EC₅₀ values for CD3ε binding from single digit to triple digit nM.This binding shift was evident in multispecific, activatable antibodiesthat utilize two versions of SP34 scFv (namely the scFv in CI005, CI023,CI024 versus the scFv in CI015, CI011, CI010). In addition, EGFR maskingis still effective in the multispecific, activatable antibodies that aremasked for both EGFR and CD3 binding.

Example 8 Cytotoxicity of Dually Masked Multispecific, ActivatableAntibodies to EGFR+ HT-29 Cells and CD3ε+ Jurkat Cells

To determine if the CD3ε masking peptides described in the previousexamples could function in the context of an EGFR masked multispecific,activatable antibody, a cytotoxicity assay was performed.

Human PBMCs were purchased as frozen aliquots (HemaCare, CatalogPB009C-1) or isolated from a buffy coat obtained from a healthy donor(Stanford Blood Center) via Ficoll density gradient centrifugation(REFERENCE=Isolation of mononuclear cells and granulocytes from humanblood. Bøyum, A., Scand. J., Clin. Lab. Invest. 21 Suppl, 97, 77-89(1968)). Human PBMCs were co-cultured in RPMI-1640+glutamax, 5% heatinactivated human serum (Sigma, Catalog H3667), and 100 U/ml penicillin,and 100 μg/ml streptomycin in a white wall 96 well plates withEGFR-expressing HT29-luc2 cells at a PBMC to HT29-luc ratio of 10:1.Titrations of the following multispecific antibodies and multispecific,activatable antibodies were tested: CI005 (EGFR/hCD3), CI023(M-EGFR/M-hCD3), CI024 (M-EGFR/M-hCD3), CI011 (M-EGFR/M-hCD3), CI010(M-EGFR/M-hCD3). In addition, EGFR antibody (CI012) and isotype wereused as negative controls at a single concentration. The total volumeafter addition of effector cells, target cells, and antibody was 150 μL.After 48 hours, the luminogenic peptide substrate of the CytoTox-Glo™Cytotoxicity Assay (Promega) was added directly to the plates to measurereleased protease activity. Luminescence was measured 50 min aftersubstrate addition on the Infinite M200 Pro (Tecan). Results wereexpressed in luminescence after background subtraction of untreatedvalues and plotted in Prism with curve fitting analysis log(agonist) vs.response (three parameters).

FIGS. 16A and 16B demonstrate that the EGFR and CD3ε maskedmultispecific, activatable antibodies shift the cytotoxicity EC₅₀relative to an unmasked control. This shift was evident inmultispecific, activatable antibodies that utilize two versions of SP34scFv, namely the scFv in CI023 and CI024 versus the scFv in CI011 andCI010.

Example 9 Binding of Singly and Dually Masked Multispecific, ActivatableAntibodies to EGFR+ HT-29 Cells and CD3ε+ Jurkat Cells

To determine if the CD3ε and EGFR masks could attenuate binding in amultispecific, activatable antibody, a flow cytometry assay wasperformed. Binding to CD3ε+ Jurkat cells and EGFR+ HT-29 cells wasperformed as previously described on the following multispecificantibody and multispecific, activatable antibodies: CI005 (EGFR/hCD3),CI007 (M-EGFR/hCD3), CI008 (EGFR/M1-hCD3), CI009 (EGFR/M2-hCD3), CI010(M-EGFR/M1-hCD3), and CI011 (M-EGFR/M2-hCD3).

FIGS. 17A and 17B demonstrate that the EGFR mask can shift the EC₅₀ ofEGFR binding in the presence (CI010, CI011) and absence (CI007) of theCD3ε masks. Both CD3ε masks can shift the EC₅₀ of CD3ε binding in thepresence (CI010, CI011) and absence (CI008, C009) of the EGFR mask.

Example 10 EGFR Dependent Cytotoxicity of Singly and Dually MaskedMultispecific, Activatable Antibodies

To determine if the CD3ε and EGFR masks could attenuate cell killing ina multispecific, activatable antibody, a cytotoxicity assay wasperformed. EGFR dependent cell killing was evaluated by co-culturingPBMCs and EGFR+ HT-29 luc cells as previously described. EGFRindependent cell killing was evaluated by co-culturing PBMCs with theEGFR negative U266 cell line (ATCC, Catalog TIB-196). U266 cells werecultured according to ATCC guidelines in RPMI-1640+glutamax, 10% HeatInactivated-Fetal Bovine Serum, 100 U/ml penicillin, and 100 μg/mlstreptomycin. Titrations of the following multispecific antibody andmultispecific, activatable antibodies were tested: CI005 (EGFR/hCD3),CI007 (M-EGFR/hCD3), CI009 (EGFR/M2-hCD3), CI011 (M-EGFR/M2-hCD3).Co-culture conditions and quantification of cytotoxicity is described inthe previous example.

FIGS. 18A and 18B demonstrate that killing of EGFR+ HT29-luc2 cells ismaximally attenuated by masking both EGFR and CD3 binding in themultispecific, activatable antibody. In contrast, killing of EGFRnegative U266 cells was undetectable in identical co-culture conditions.

Example 11 EGFR Dependent Primary T Cell Activation of Singly and DuallyMasked Multispecific, Activatable Antibodies

To determine if the CD3ε and EGFR masks could attenuate primary T cellactivation in a multispecific, activatable antibody, a flow cytometryassay was performed. PBMC and EGFR+ HT-29 luc2 co-culture conditions aredescribed in the previous example. After a 24 hour incubation, theculture was transferred to a U-bottom plate and the remaining cells weredetached with 0.25% trypsin (Life Technologies, Catalog 25200-056).Trypsin activity was quenched by adding 3 volumes of FACS Buffer, andthe cell suspension was transferred to the same U-bottom plate. Afterharvesting, the cells were resuspended in 50 μL of either an anti-CD69PE/Anti-CD8 APC-Vio770 cocktail (anti-CD69 PE, BD Biosciences, Catalog555531; anti-CD8 APC-Vio770, Miltenyi Biotec, Catalog 130-096-561),APC-Vio770 isotype control (Miltenyi Biotec, Catalog 130-099-637), or PEisotype control (BD BioSciences, Catalog 340761). All antibodies wereused at the manufacturer's recommended concentrations. Cells werestained for 1 h at 4° C. with shaking, harvested and re-suspended in afinal volume of 100 μL FACS Buffer with 2.5 μg/ml 7-AAD. Samples werecollected on a MACSQuant® Analyzer 10 (Miltenyi) and activation wasquantified in FlowJo V10 (Treestar) as the percentage of CD8+ T cellswith expression of CD69 above the PE isotype control. EC₅₀ values werecalculated in GraphPad Prism 6 by curve fitting the data to log(agonist)vs. response (three parameters).

FIG. 19 demonstrates that activation of primary CD8+ T cells ismaximally attenuated by masking both EGFR and CD3 binding in themultispecific, activatable antibody.

Example 12 EGFR Independent Primary T Cell Activation of Singly andDually Masked Multispecific, Activatable Antibodies

To determine if the CD3ε masks could attenuate EGFR independentactivation in a multispecific, activatable antibody, a flow cytometryassay was performed. PBMC and EGFR negative U266 co-culture conditionsare described in the previous example. Activation of primary CD8+ Tcells was assessed by CD69 staining as previously described. Thefollowing multispecific antibody and multispecific, activatableantibodies were evaluated: CI005 (EGFR/hCD3), CI007 (M-EGFR/hCD3), CI009(EGFR/M2-hCD3), CI011 (M-EGFR/M2-hCD3).

FIGS. 20A and 20B demonstrate that target-independent T cell activationby the unmasked bispecific antibody is only very minimal and that thislow degree of activity is abrogated upon masking of the anti-CD3 moiety.

Example 13 uPA Digest of Multispecific, Activatable Antibodies RestoresBinding to EGFR+ HT-29 Luc2 Cells and CD3+ Jurkat Cells and CytotoxicityAgainst EGFR+ HT-29 Luc2 Cells

Activation of the multispecific activatable antibody was conducted asfollows: multispecific activatable antibodies in PBS was cleaved by theaddition of uPA (R&D Systems, Catalog 1310-SE) to a final concentrationof about 1 μM. The digest was incubated at 37° C. overnight, andcleavage was confirmed by removing an aliquot for capillaryelectrophoresis analysis (GX-II Capillary Electrophoresis, Perkin Elmer)or SDS-PAGE. Protease and the cleaved masking moiety were removed byProtein A purification. Briefly, the digested sample was diluted to 2 mlwith PBS and loaded onto equilibrated MabSelect SuRe™ beads (GEHealthcare Life Sciences, Product 11-0026-01 AD). Beads were washed with5 column volumes (CV) of 1×PBS, followed by 5 CV of 5×PBS supplementedwith 5% isopropyl alcohol (IPA), and finally with 5 CV of 1×PBS.Antibody was eluted with 10 CV of 0.1 M Glycine, pH 3.0, and fractionswere neutralized with 1 M Tris, pH 8.0, pooled, concentrated and bufferexchanged into PBS. Binding to CD3ε+ Jurkat cells and EGFR+ HT-29 cellsand cell killing against HT-29 luc2 cells was performed as previouslydescribed.

FIGS. 21A-21D demonstrate that uPA activation restores binding of themultispecific, activatable CI011 (M-EGFR/M-hCD3) antibody to both EGFR+HT29-luc2 cells and CD3+ Jurkat cells. In addition, cell killing of allmultispecific, activatable antibodies is restored by uPA activation.

Example 14 Dually Masked Multispecific, Activatable AntibodiesIncorporating Different Substrates

To determine if different substrates impact EGFR dependent killing ofdually masked, multispecific, activatable antibodies, a cytotoxicityassay was performed. Cell killing against HT-29 luc2 cells was performedas previously described.

FIG. 22 demonstrates that dually masked multispecific, activatableantibodies that contain different substrates shift EC₅₀ cytotoxicity.

Example 15 Multispecific Antibodies and Multispecific ActivatableAntibodies of the Embodiments Prevent HT-29Luc2 Tumor Growth in Mice

In this example, multispecific antibodies and multispecific activatableantibodies targeting EGFR and CD3ε were analyzed for ability to preventgrowth of HT-29Luc2 xenograft tumors in immune-deficient NOD-scid mice.

The human colon cancer cell line HT-29Luc2 was obtained from PerkinElmer, Inc., Waltham, Mass. (formerly Caliper Life Sciences, Inc.). Thefresh, human PBMCs and PBMCs enriched for T cells were obtained from ahuman donor through Hemacare, Inc., Van Nuys, Calif. and shippedovernight to CytomX. The HT-29Luc2 cells were implanted subcutaneouslyalong with human PBMCs or fresh PBMCs enriched for T cells into theright flank of NOD scid (NOD.CB17-Prkdc^(scid)/J) mice (JacksonLaboratory, Bar Harbor, Me.). The HT-29Luc2 cells were grown in McCoy's5a medium (ATCC, Manassas, Va.), with 10% fetal bovine serum(Gibco-brand, ThermoFisher, Inc. Waltham, Mass.) at 37° C. in anatmosphere of 5% CO₂ in air. The tumor cells were routinely sub-culturedtwice weekly. Cells were harvested during the logarithmic growth periodand kept at ambient temperature for tumor induction.

In one prevention study, mice were inoculated subcutaneously with 100 μLof a cell suspension containing 2×10⁶ tumor cells with or without 2×10⁶fresh, unstimulated PBMC (Day 0). On the same day, mice received a firstdose of test or control article by intravenous injection (Day 0). Tumorvolume was measured twice weekly in two dimensions using a digitalcaliper, and the volume was expressed in mm³ using the formula: V=0.5a×b² where a and b are the long and short diameters of the tumor,respectively.

The mice were grouped and dosed as set forth in Table 13.

TABLE 13 Groups and doses for HT-29Luc2 xenograft study. Dose GroupCount Tumor PBMC Treatment (mg/kg) Schedule 1 8 HT-29Luc2 — PBS — q7dx2,iv 2 8 HT-29Luc2 Donor A PBS — q7dx2, iv 3 8 HT-29Luc2 Donor A CI0050.10 q7dx2, iv 4 8 HT-29Luc2 Donor A CI059 0.5*  qdx8, iv (BiTE) *BiTEwas dosed at 0.5 mg/kg/day.

FIG. 23, which plots tumor volume versus days post initial dose,demonstrated that multispecific antibody CI005 at 0.1 mg/kg dosed twice7 days apart and CI059 (BiTE) at 0.5 mg/kg/day for 8 consecutive daysprevented the growth of HT-29Luc2 xenograft tumors.

In a separate study, multispecific antibody CI048 and multispecificactivatable antibody CI011 targeting EGFR and CD3ε were tested for theability to prevent or inhibit the growth of HT-29Luc2 xenograft tumorusing a method similar to that described above. Groups and doses are setforth in Table 14. The effector cells in this study were fresh,unstimulated CD3+ T cells enriched from PBMCs, and were co-injected in a1:1 ratio of effector cell to tumor cell.

TABLE 14 Groups and doses for HT-29Luc2 xenograft prevention study.Route/volume Effector of Dose Group Count Tumor cells AdministrationTreatment (mg/kg) Schedule 1 8 HT-29Luc2 Donor A SC, co- PBS — d0, d7 29 HT-29Luc2 Donor A administration CI048 0.10 d0, d7 3 10 HT-29Luc2Donor A with tumor on CI011 0.10 d0, d7 d0 with IV administration oftreatment 5 ml/kg

FIG. 24, which plots tumor volume versus days post initial dose,demonstrated that CI048 multispecific antibody limited establishment ofHT-29Luc2 xenograft tumors and CI011 multispecific activatable antibodyinhibited the growth of HT-29Luc2 xenograft tumors. The difference ineffect between PBS and CI011 on Day 26 is statistically significant(p<0.05).

Example 16 Multispecific Antibodies and Multispecific ActivatableAntibodies of the Embodiments Induced Regression of EstablishedHT-29Luc2 Tumors in Mice

In this example, multispecific antibodies and multispecific activatableantibodies targeting EGFR and CD3ε were analyzed for the ability toproduce regression or reduced growth of established HT-29Luc2 xenografttumors in human T-cell engrafted NSG mice.

The human colon cancer cell line HT-29Luc2 was obtained from PerkinElmer, Inc., Waltham, Mass. (formerly Caliper Life Sciences, Inc.) andcultured as described in Example 15. Purified, frozen human PBMCs wereobtained from a human donor through Hemacare, Inc., Van Nuys, Calif. NSG(NOD.Cg-Prkdcscid Il2rg^(tm1Wj1)/SzJ) mice were obtained from TheJackson Laboratories, Bar Harbor, Me.

In preparation for the tumor intervention study, each mouse wasinoculated subcutaneously at the right flank with 2×10⁶ HT-29Luc2 cellsin 100 μL RPMI+Glutamax, serum-free medium. Previously frozen PBMCs froma single donor (“A”) were analyzed to determine the percent CD3+ Tcells. This value was used to calculate the total number of PBMCs neededto achieve a CD3+ T cell to tumor cell ratio of 2:1. This total numberof PBMCs (10.2 million) was injected into the peritoneum of each mouseon the day of tumor cell implantation.

Blood (˜100 μl) was collected from all mice 11 days post PBMCinoculation to evaluate extent of human T cell engraftment within eachmouse by flow cytometry. Whole blood (anti-coagulant, lithium heparin)was assayed for presence of human CD3+ with percent CD4+ and CD8+ Tcells.

Tumor volume and body weights were measured twice weekly and tumorvolumes calculated as described in Example 15. 13 days after tumorinoculation, mice with adequate T cell engraftment and with tumors inthe target size range (50-100 mm³; average ˜70 mm³) were randomized intogroups of 8. The following day, mice assigned to treatment groups weredosed IV (5 mL/kg) according to Table 15.

TABLE 15 Groups and doses for dose dependent HT-29Luc2 xenograft study.Route of Route of Administration Administration Dose Group Count Tumor¹Effector cells^(2, 3) Effector cells compound Treatment (mg/kg) Schedule1 8 HT-29Luc2 10.2 million/mouse IP IV PBS — d1, d8, 2 8 HT-29Luc2 10.2million/mouse CI048 0.003 d15 3 8 HT-29Luc2 10.2 million/mouse CI0480.010 4 8 HT-29Luc2 10.2 million/mouse CI048 0.030 5 8 HT-29Luc2 10.2million/mouse CI048 0.10 6 8 HT-29Luc2 10.2 million/mouse CI048 0.30 7 8HT-29Luc2 10.2 million/mouse CI040 0.10 ¹2 million HT29-Luc2 cells wereimplanted. ²PBMC number were normalized to CD3+ number and with a ratioof 1:2 tumor:CD3+ T cell. ³frozen PBMCs were allowed to engraft withoutpre-stimulation for ^(~)13 days.

FIG. 25, which plots tumor volume versus days post initial treatmentdose, demonstrated a dose-dependent effect of CI048 multispecificantibody on the growth of HT-29Luc2 xenograft tumors. The mostefficacious dose of CI048 was 0.3 mg/kg, resulting in sustained,complete tumor regression in 100% mice. CI048 at 0.1 mg/kg was alsohighly efficacious, with a majority of mice exhibiting tumor regression.

FIG. 26, which plots tumor volume versus days post initial treatmentdose, demonstrated that CI048 multispecific antibody at 0.3 mg/kgeliminated HT-29Luc2 xenograft tumors and the CI040 multispecificactivatable antibody at 0.1 mg/kg is less efficacious than CI048 butlimited the growth of HT-29Luc2 xenograft tumors relative to a PBScontrol.

Two additional efficacy studies were set up using unassigned animalsfrom the same pool of HT-29Luc2 tumor-bearing, human T cell engraftedNSG mice. The first study was initiated 18 days post tumor implantationwith an average starting tumor volume of 107 mm³. This study comparedthe activities of CI048 multispecific antibody and CI040 multispecificactivatable antibody. The mice were grouped and dosed as set forth inTable 16, Groups 1-4. The second study was initiated 26 days post tumorimplantation with an average starting tumor volume of 190 mm³. Thisstudy evaluated the efficacy of CI011 multispecific activatableantibody. The mice were grouped and dosed IV as set forth in Table 16,Groups 5 and 6.

TABLE 16 Groups and doses for additional dose dependent HT-29Luc2xenograft add-on studies. Route of Route of AdministrationAdministration Dose Group Count Tumor¹ Effector cells^(2, 3) Effectorcells compound Treatment (mg/kg) Schedule 1 7 HT-29Luc2 10.2million/mouse IP IV PBS — d1, d8, 2 7 HT-29Luc2 10.2 million/mouse CI0480.30 d15, d22 3 7 HT-29Luc2 10.2 million/mouse CI040 0.30 4 7 HT-29Luc210.2 million/mouse CI040 1.0 6 4 HT-29Luc2 10.2 million/mouse PBS — d1,d8 7 4 HT-29Luc2 10.2 million/mouse CI011 1.0 ¹2 million HT-29Luc2 cellswere implanted. ²PBMC number were normalized to CD3+ number and with aratio of 1:2 tumor:CD3+ T cell. ³frozen PBMCs were allowed to engraftwithout pre-stimulation for ^(~)19-21 days.

FIG. 27, which plots tumor volume versus days post initial treatmentdose, demonstrated that the CI040 multispecific activatable antibody at1.0 mg/kg and the CI048 multispecific antibody at 0.3 mg/kg producedcomplete tumor regression in 100% of treated mice. Tumor regression wasseen in a majority of mice dosed with 0.3 mg/kg CI040.

FIG. 28, which plots tumor volume versus days post initial treatmentdose, demonstrated CI011 multispecific activatable antibody at 1.0 mg/kgsignificantly inhibited HT-29Luc2 xenograft tumor growth relative to avehicle control (p<0.05).

Example 17 Maximum Tolerated Dose (MTD) Determination for aMultispecific Antibody and Multispecific Activatable Antibodies inCynomolgus Monkeys

In this example, an escalating dose study design was used to establishthe maximum tolerated dose of CI048 EGFRxCD3ε multispecific antibody andCI011 and CI040, two multispecific activatable antibodies following asingle IV bolus administration to naïve male cynomolgus monkeys. Themonkeys were of Cambodian origin and ranged in weight from 2.5 to 4 kg.CI011 and CI040 varied only in the amino acid sequence of theprotease-cleavable site. CI048, CI011 and CI040 were each administeredto one animal per dose level beginning at 0.1 micrograms/kg (μg/kg) forCI048 and at 60 μg/kg for CI011 and CI040. The dose was escalated in 2to 10-fold increments until a MTD for each article was reached orexceeded. One additional animal was then used to confirm the MTD. Eachstudy animal was dosed once and monitored for a minimum of 7 days.Tolerability was evaluated based on clinical signs, body weight, foodconsumption and laboratory analyses that included serum chemistry andhematology. Blood was collected for standard serum chemistry andhematology analysis once during acclimation and at 48 h and 7 days postdose.

Abnormal clinical signs including emesis, hunching and reduced foodintake were observed with the unmasked CI048 multispecific antibody atdoses of 10 μg/kg and above, and with CI011 or CI040 multispecificactivatable antibodies at doses ≧200 μg/kg. At the MTDs of 20 μg/kg forCI048, ≧200 μg/kg for CI040 and ≧600 μg/kg for CI011, these findings,when present, were transient and generally confined to the 48 hrpost-dose period. Serum chemistry findings at the MTD included mild tomoderate elevations of alanine transaminase (ALT, a liver enzyme markerfound in serum upon liver cell lysis) at 48 hr that were fully reversedby Day 8. A single animal dosed with CI048 at 60 μg/kg, demonstratedmore severe and protracted clinical signs, correlating withsubstantially elevated ALT, AST, bilirubin and blood urea nitrogen at 48hr post dose. This animal also had marked increases in serum cytokinesincluding IL-6, IFN-gamma and TNF-alpha at 1 or 4 hr post dose, and amassively expanded lymphocyte population on Day 8. The findings of liverand kidney injury at CI048 doses exceeding the MTD are similar to thosenoted by Lutterbuese and co-workers in a study with an EGFR×CD3bispecific molecule (PNAS 2010, vol. 107(28) 12605-12610) and may beattributable to T cell-mediated tissue injury.

FIG. 29 depicts 48 hr post-dose serum concentration of alanineaminotransferase (ALT) as a function of test article dose. A markedincrease in ALT was noted at 60 μg/kg CI048 multispecific antibody.Re-dosing of CI048 at 20 μg/kg established the tolerated dose at 20μg/kg. Administration of CI011 and CI040 multispecific activatableantibodies at doses of 600 μg/kg resulted in serum ALT levels that werecomparable to serum ALT concentrations associated with 10-20 μg/kgCI048.

Example 18 Binding of Dually Masked Multispecific Activatable Antibodiesto EGFR+HT-29 Cells and Mouse CD3ε+ TK1 Cells

To determine if mouse CD3ε and EGFR masks could attenuate binding in amultispecific activatable antibody, a flow cytometry assay wasperformed. Mouse CD3ε+ TK1 cells (ATCC, Catalog CRL-2396) were culturedin RPMI-1640+glutamax, 10% Heat Inactivated-Fetal Bovine Serum, 100 U/mlpenicillin, and 100 μg/ml streptomycin according to ATCC guidelines.Binding to mouse CD3ε+ TK1 cells and EGFR+ HT-29 cells was performed aspreviously described on the following multispecific antibody andmultispecific activatable antibodies: CI052 (EGFR/mCD3), CI053(M-EGFR/M01-mCD3), CI054 (M-EGFR/M02-mCD3), CI055 (M-EGFR/M03-mCD3),CI056 (M-EGFR/M04-mCD3), CI049 (M-EGFR/M05-mCD3), CI050(M-EGFR/M06-mCD3), CI051 (M-EGFR/M07-mCD3), CI057 (M-EGFR/M08-mCD3), andCI058 (M-EGFR/M09/mCD3). FIG. 30 demonstrates that incorporation of themouse CD3ε masks into the EGFR masked multispecific activatable antibodyshifted the EC₅₀ values for mouse CD3ε binding while retaining effectiveEGFR masking.

Example 19 Characterization of IL6R Targeting Dually MaskedMultispecific Activatable Antibodies

To determine if the CD3ε and IL6R masks of a multispecific activatableantibody could attenuate cell killing, a cytotoxicity assay wasperformed. Cell killing was evaluated by co-culturing purified humanCD8+ cells and IL6R+Molp-8 cells in a 6 to 1 ratio. Human CD8+ T cellswere isolated from frozen PBMCs by negative selection using theDynabeads® Untouched™ Human CD8 T Cells Kit (Life Technologies, Catalog11348D). Molp-8 cells (DSMZ, Catalog ACC 569) were cultured according toDSMZ guidelines in RPMI-1640+glutamax, 20% Heat Inactivated-Fetal BovineSerum, 100 U/ml penicillin, and 100 μg/ml streptomycin. Titrations ofthe following multispecific antibody and multispecific, activatableantibodies were tested: CI026 (IL6R/hCD3), CI027 (IL6R/M-hCD3), CI036(M-IL6R/hCD3) and CI029 (M-IL6R/M-hCD3). Co-culture conditions andquantification of cytotoxicity is described in the previous example.

FIG. 31 demonstrates that killing of IL6R+Molp-8 cells is maximallyattenuated by masking both IL6R and CD3 binding in the multispecific,activatable antibody.

Other Embodiments

While the invention has been described in conjunction with the detaileddescription thereof, the foregoing description is intended to illustrateand not limit the scope of the disclosure, which is defined by the scopeof the appended claims. Other aspects, advantages, and modifications arewithin the scope of the following claims.

What is claimed is:
 1. An isolated single chain variable antibodyfragment that binds the epsilon chain of CD3 (CD3ε) comprising avariable heavy chain complementarity determining region 1 (VH CDR1)comprising the amino acid sequence TYAMN (SEQ ID NO: 53); a variableheavy chain complementarity determining region 2 (VH CD2) comprising theamino acid sequence RIRSKYNNYATYYADSVKD (SEQ ID NO: 54); a variableheavy chain complementarity determining region 3 (VH CDR3) comprisingthe amino acid sequence HGNFGNSYVSWFAY (SEQ ID NO: 55), a variable lightchain complementarity determining region 1 (VL CDR1) comprising theamino acid sequence RSSTGAVTTSNYAN (SEQ ID NO: 56); a variable lightchain complementarity determining region 2 (VL CDR2) comprising theamino acid sequence GTNKRAP (SEQ ID NO: 57); and a variable light chaincomplementarity determining region 3 (VL CDR3) comprising the amino acidsequence ALWYSNLWV (SEQ ID NO: 58).
 2. The isolated single chainvariable antibody fragment of claim 1, wherein the AB comprises avariable heavy chain (Hv) comprising the amino acid sequence of SEQ IDNO: 4 and a variable light chain (Lv) comprising the amino acid sequenceof SEQ ID NO:
 2. 3. The isolated single chain variable antibody fragmentof claim 1, wherein the scFv fragment comprises the amino acid sequenceof SEQ ID NO: 6 or the amino acid sequence of SEQ ID NO:
 30. 4. Theisolated single chain variable antibody fragment of claim 1, wherein theAB comprises the amino acid sequence of SEQ ID NO:
 587. 5. The isolatedsingle chain variable antibody fragment of claim 1, wherein the ABcomprises the amino acid sequence of SEQ ID NO:
 588. 6. The isolatedsingle chain variable antibody fragment of claim 1 comprising an agentconjugated to the antibody.
 7. The isolated single chain variableantibody fragment of claim 6, wherein the agent is a therapeutic agent,an antineoplastic agent, a toxin or fragment thereof, a detectablemoiety or a diagnostic agent.
 8. The isolated single chain variableantibody fragment of claim 6, wherein the agent is conjugated to theantibody via a linker.
 9. A pharmaceutical composition comprising anantibody as in any of claims 1 to 8 and a carrier.
 10. A pharmaceuticalcomposition comprising an antibody as in any of claims 1 to 5 and acarrier.
 11. An activatable antibody that in an activated state bindsthe epsilon chain of CD3 (CD3ε), the activatable antibody comprising: anantibody or an antigen binding fragment thereof (AB) that specificallybinds to CD3ε; a masking moiety (MM) that inhibits the binding of the ABto CD3ε when the activatable antibody is in an uncleaved state; and acleavable moiety (CM) coupled to the AB, wherein the CM is a polypeptidethat functions as a substrate for a protease.
 12. The activatableantibody of claim 11, wherein the MM has a dissociation constant forbinding to the AB that is selected from the group consisting of: greaterthan the dissociation constant of the AB to CD3ε; equivalent to thedissociation constant of the AB to CD3ε; and less than the dissociationconstant of the AB to CD3ε.
 13. The activatable antibody of claim 11,wherein the MM does not interfere or compete with the AB for binding toCD3ε when the activatable antibody is in a cleaved state.
 14. Theactivatable antibody of claim 11, wherein the MM is a polypeptide of nomore than 40 amino acids in length.
 15. The activatable antibody ofclaim 11, wherein the MM polypeptide sequence is different from that ofCD3ε and wherein the MM polypeptide sequence is no more than 50%identical to any natural binding partner of the AB.
 16. The activatableantibody of claim 11, wherein the MM comprises a sequence selected fromthe group consisting of the sequences shown in Table 7 or Table
 8. 17.The activatable antibody of claim 11, wherein the protease is producedby a tumor that is in proximity to cells that express CD3ε in a tissueand/or produced by a tumor that is co-localized with CD3ε in a tissue,and wherein the protease cleaves the CM in the activatable antibody whenthe activatable antibody is exposed to the protease.
 18. The activatableantibody of claim 11, wherein the CM is a polypeptide of up to 15 aminoacids in length.
 19. The activatable antibody of claim 11, wherein theCM is a substrate for a protease selected from the group consisting ofthe proteases shown in Table 3 or wherein the CM comprises an amino acidsequence selected from the group consisting of SEQ ID NO: 67-86,321-341, and 896-926.
 20. The activatable antibody of claim 11, whereinthe antigen binding fragment thereof is selected from the groupconsisting of a Fab fragment, a F(ab′)₂ fragment, a scFv, a scab, a dAb,a single domain heavy chain antibody, and a single domain light chainantibody.
 21. The activatable antibody of claim 11, wherein theactivatable antibody in the uncleaved state has the structuralarrangement from N-terminus to C-terminus as follows: MM-CM-AB orAB-CM-MM.
 22. The activatable antibody of claim 11, wherein theactivatable antibody comprises a linking peptide between the MM and theCM.
 23. The activatable antibody of claim 11, wherein the activatableantibody comprises a linking peptide between the CM and the AB.
 24. Theactivatable antibody of claim 11, wherein the activatable antibodycomprises a first linking peptide (LP1) and a second linking peptide(LP2), and wherein the activatable antibody in the uncleaved state hasthe structural arrangement from N-terminus to C-terminus as follows:MM-LP1-CM-LP2-AB or AB-LP2-CM-LP1-MM.
 25. The activatable antibody ofclaim 24, wherein the two linking peptides are not identical to eachother.
 26. The activatable antibody of claim 24, wherein each of LP1 andLP2 is a peptide of about 1 to 20 amino acids in length.
 27. Theactivatable antibody as in any of claims 11 to 26, wherein the ABcomprises a VH CDR1 comprising the amino acid sequence TYAMN (SEQ ID NO:53); a VH CD2 comprising the amino acid sequence RIRSKYNNYATYYADSVKD(SEQ ID NO: 54); a VH CDR3 comprising the amino acid sequenceHGNFGNSYVSWFAY (SEQ ID NO: 55), a VL CDR1 comprising the amino acidsequence RSSTGAVTTSNYAN (SEQ ID NO: 56); a VL CDR2 comprising the aminoacid sequence GTNKRAP (SEQ ID NO: 57); and a VL CDR3 comprising theamino acid sequence ALWYSNLWV (SEQ ID NO: 58).
 28. The activatableantibody as in any of claims 11 to 26, wherein the AB comprises theamino acid sequence of SEQ ID NO:
 587. 29. The activatable antibody asin any of claims 11 to 26, wherein the AB comprises the amino acidsequence of SEQ ID NO:
 588. 30. The activatable antibody as in any ofclaims 11 to 26, wherein the activatable antibody comprises a heavychain sequence selected from the group consisting of SEQ ID NOs: 446,452, 454, 456, 460, 462, 464, 466, 470, 472, 476, 478, 480, 486, 488,490, 492, 494, 496, 498, 500, 502, 504, 510, 512, 514, 518, 524, 526,530, 532, 534, 536, 538, 540, 542, 544, and
 546. 31. The activatableantibody as in any of claims 11 to 26, wherein the activatable antibodycomprises a light chain sequence selected from the group consisting ofSEQ ID NOs: 448, 450, 458, 468, 474, 482, 484, 508, 516, and
 520. 32.The activatable antibody as in any of claims 11 to 26, wherein theactivatable antibody comprises a heavy chain sequence selected from thegroup consisting of SEQ ID NOs: 446, 452, 454, 456, 460, 462, 464, 466,470, 472, 476, 478, 480, 486, 488, 490, 492, 494, 496, 498, 500, 502,504, 510, 512, 514, 518, 524, 526, 530, 532, 534, 536, 538, 540, 542,544, and 546; and a light chain sequence selected from the groupconsisting of SEQ ID NOs: 448, 450, 458, 468, 474, 482, 484, 508, 516,and
 520. 33. The activatable antibody as in any of claims 11 to 26,wherein the activatable antibody comprises the amino acid sequence ofSEQ ID NO:
 506. 34. The activatable antibody as in any of claims 11 to33 comprising an agent conjugated to the AB.
 35. The activatableantibody of claim 34, wherein the agent is a therapeutic agent, anantineoplastic agent, a toxin or fragment thereof, a detectable moietyor a diagnostic agent.
 36. The activatable antibody of claim 34, whereinthe agent is conjugated to the AB via a linker.
 37. The activatableantibody of claim 11, wherein the antibody or antigen-binding fragmentthereof comprises a variable heavy chain (Hv) comprising the amino acidsequence of SEQ ID NO: 4 and a variable light chain (Lv) comprising theamino acid sequence of SEQ ID NO:
 2. 38. The activatable antibody ofclaim 11, wherein the antibody or antigen-binding fragment thereofcomprises a scFv fragment.
 39. The activatable antibody of claim 38,wherein the scFv fragment comprises the amino acid sequence of SEQ IDNO: 6 or the amino acid sequence of SEQ ID NO:
 30. 40. A pharmaceuticalcomposition comprising an activatable antibody as in any of claim 11 to33 or 37-39 and a carrier.
 41. A pharmaceutical composition comprisingan activatable antibody as in any of claims 11 to 39 and a carrier. 42.An isolated nucleic acid molecule encoding at least a portion of theisolated single chain variable antibody as in any of claims 1 to
 5. 43.An isolated nucleic acid molecule encoding at least a portion of theactivatable antibody as in any of claim 11-33 or 37-39.
 44. A vectorcomprising the isolated nucleic acid molecule of claim 42 or claim 43.45. A method of producing an isolated single chain variable antibody oran activatable antibody by culturing a cell under conditions that leadto expression of the antibody or the activatable antibody, wherein thecell comprises the nucleic acid molecule of claim 42 or claim
 43. 46. Amethod of manufacturing an activatable antibody that binds the epsilonchain of CD3 (CD3ε) in an activated state, the method comprising: (a)culturing a cell comprising a nucleic acid construct that encodes theactivatable antibody under conditions that lead to expression of theactivatable antibody, wherein the activatable antibody comprises amasking moiety (MM), a cleavable moiety (CM), and an antibody or anantigen binding fragment thereof (AB) that specifically binds theepsilon chain of CD3 (CD3ε), and (b) recovering the activatableantibody.
 47. The method of claim 46, wherein the CM is a polypeptidethat functions as a substrate for a protease.
 48. The method of claim46, wherein the CM is positioned in the activatable antibody such that,when the activatable antibody is in an uncleaved state, the MMinterferes with specific binding of the AB to CD3ε and when theactivatable antibody is in a cleaved state the MM does not interfere orcompete with specific binding of the AB to CD3ε.
 49. An isolatedmultispecific antibody that binds the epsilon chain of CD3 (CD3ε) and asecond target, wherein the antibody comprises a first antibody orantigen-binding fragment thereof (AB1) that binds the epsilon chain ofCD3 (CD3ε) and a second antibody or antigen-binding fragment thereof(AB2) that binds a second target, and wherein AB1 comprises a VH CDR1comprising the amino acid sequence TYAMN (SEQ ID NO: 53); a VH CD2comprising the amino acid sequence RIRSKYNNYATYYADSVKD (SEQ ID NO: 54);a VH CDR3 comprising the amino acid sequence HGNFGNSYVSWFAY (SEQ ID NO:55), a VL CDR1 comprising the amino acid sequence RSSTGAVTTSNYAN (SEQ IDNO: 56); a VL CDR2 comprising the amino acid sequence GTNKRAP (SEQ IDNO: 57); and a VL CDR3 comprising the amino acid sequence ALWYSNLWV (SEQID NO: 58).
 50. The multispecific antibody of claim 49, wherein AB1comprises a variable heavy chain (Hv) comprising the amino acid sequenceof SEQ ID NO: 4 and a variable light chain (Lv) comprising the aminoacid sequence of SEQ ID NO:
 2. 51. The multispecific antibody of claim49, wherein the AB1 comprises a scFv fragment.
 52. The multispecificantibody of claim 51, wherein the scFv fragment comprises the amino acidsequence of SEQ ID NO: 6 or the amino acid sequence of SEQ ID NO: 30.53. The multispecific antibody of claim 49, wherein the AB comprises theamino acid sequence of SEQ ID NO:
 587. 54. The multispecific antibody ofclaim 49, wherein the AB comprises the amino acid sequence of SEQ ID NO:588.
 55. The antibody of claim 49 comprising an agent conjugated to theantibody.
 56. The multispecific antibody of claim 49, wherein the agentis a therapeutic agent, an antineoplastic agent, a toxin or fragmentthereof, a detectable moiety or a diagnostic agent.
 57. Themultispecific antibody of claim 49, wherein the agent is conjugated tothe antibody via a linker.
 58. The multispecific antibody of claim 57,wherein the linker is a cleavable linker.
 59. The multispecific antibodyof claim 57, wherein the linker is a non-cleavable linker.
 60. Themultispecific antibody of claim 49, wherein the multispecific antibodyis selected from: a multispecific antibody comprising: a cleavablemoiety (CM) coupled to the AB2, wherein the CM is a polypeptide thatfunctions as a substrate for a protease; and a masking moiety (MM) thatinhibits the binding of the AB2 to the second target when the CM is inan uncleaved state; and a multispecific antibody comprising: a cleavablemoiety (CM) coupled to the AB1, wherein the CM is a polypeptide thatfunctions as a substrate for a protease; and a masking moiety (MM) thatinhibits the binding of the AB1 to CD3ε when the CM is in an uncleavedstate.
 61. An isolated multispecific activatable antibody that binds theepsilon chain of CD3 (CD3ε) and a second target, the multispecificactivatable antibody comprising a first antibody or an antigen bindingfragment thereof (AB1) that specifically binds to CD3ε and a secondantibody or antigen-binding fragment thereof (AB2) that binds a secondtarget, wherein AB1 comprises a VH CDR1 comprising the amino acidsequence TYAMN (SEQ ID NO: 53); a VH CD2 comprising the amino acidsequence RIRSKYNNYATYYADSVKD (SEQ ID NO: 54); a VH CDR3 comprising theamino acid sequence HGNFGNSYVSWFAY (SEQ ID NO: 55), a VL CDR1 comprisingthe amino acid sequence RSSTGAVTTSNYAN (SEQ ID NO: 56); a VL CDR2comprising the amino acid sequence GTNKRAP (SEQ ID NO: 57); and a VLCDR3 comprising the amino acid sequence ALWYSNLWV (SEQ ID NO: 58); amasking moiety (MM) that inhibits the binding of the AB2 to the secondtarget when the multispecific activatable antibody is in an uncleavedstate; and a cleavable moiety (CM) coupled to the AB2, wherein the CM isa polypeptide that functions as a substrate for a protease.
 62. Themultispecific activatable antibody of claim 61, wherein the target ofAB2 is selected from the group of targets listed in Table
 1. 63. Themultispecific activatable antibody of claim 61, wherein AB2 is or isderived from an antibody selected from the group of antibodies listed inTable
 2. 64. The multispecific activatable antibody of claim 61, whereinthe antigen binding fragment thereof is selected from the groupconsisting of a Fab fragment, a F(ab′)₂ fragment, a scFv, a scAb, a dAb,a single domain heavy chain antibody, and a single domain light chainantibody.
 65. The multispecific activatable antibody of claim 61,wherein the MM has a dissociation constant for binding to the AB that isselected from the group consisting of: greater than the dissociationconstant of the AB to the target; equivalent to the dissociationconstant of the AB to CD3ε; and less than the dissociation constant ofthe AB to CD3ε.
 66. The multispecific activatable antibody of claim 61,wherein the MM does not interfere or compete with the AB for binding tothe target when the multispecific activatable antibody is in a cleavedstate.
 67. The multispecific activatable antibody of claim 61, whereinthe MM is a polypeptide of no more than 40 amino acids in length. 68.The multispecific activatable antibody of claim 61, wherein the MMpolypeptide sequence is different from that of the target of AB2, andwherein the MM polypeptide sequence is no more than 50% identical to anynatural binding partner of AB2.
 69. The multispecific activatableantibody of claim 61, wherein the CM is a polypeptide of up to 15 aminoacids in length.
 70. The multispecific activatable antibody of claim 61,wherein the protease is produced by a tumor that is in proximity tocells that express CD3ε in a tissue and/or produced by a tumor that isco-localized with the target of AB2 in a tissue, and wherein theprotease cleaves the CM in the multispecific activatable antibody whenthe multispecific activatable antibody is exposed to the protease. 71.The multispecific activatable antibody of claim 61, wherein themultispecific activatable antibody comprises a linking peptide betweenthe MM and the CM.
 72. The multispecific activatable antibody of claim61, wherein the multispecific activatable antibody comprises a linkingpeptide between the CM and the AB.
 73. The multispecific activatableantibody as in any of claims 61 to 72, wherein the multispecificactivatable antibody comprises a first linking peptide (LP1) and asecond linking peptide (LP2), and wherein the multispecific activatableantibody in an uncleaved state has the structural arrangement fromN-terminus to C-terminus as follows: MM-LP1-CM-LP2-AB orAB-LP2-CM-LP1-MM.
 74. The multispecific activatable antibody of claim73, wherein the two linking peptides are not identical to each other.75. The multispecific activatable antibody of claim 73, wherein each ofLP1 and LP2 is a peptide of about 1 to 20 amino acids in length.
 76. Themultispecific activatable antibody of claim 73, wherein at least one ofLP1 or LP2 comprises an amino acid sequence selected from the groupconsisting of (GS)_(n), (GGS)_(n), (GSGGS)_(n) (SEQ ID NO: 59) and(GGGS)_(n) (SEQ ID NO: 60), GGSG (SEQ ID NO: 61), GGSGG (SEQ ID NO: 62),GSGSG (SEQ ID NO: 63), GSGGG (SEQ ID NO: 64), GGGSG (SEQ ID NO: 65), andGSSSG (SEQ ID NO: 66), where n is an integer of at least one.
 77. Themultispecific activatable antibody of claim 61, wherein the CM is asubstrate for a protease selected from the group consisting of theproteases shown in Table 3 or wherein the CM comprises an amino acidsequence selected from the group consisting of SEQ ID NO: 67-86,321-341, and 896-926.
 78. A multispecific activatable antibody that inan activated state binds the epsilon chain of CD3 (CD3ε) and a secondtarget, the multispecific activatable antibody comprising: at least afirst antibody or an antigen binding fragment thereof (AB1) thatspecifically binds CD3ε; a second antibody or an antigen bindingfragment thereof (AB2) that specifically binds a second target orepitope; at least a first masking moiety (MM1) that inhibits the bindingof the AB1 to CD3ε when the multispecific activatable antibody is in anuncleaved state; and at least a first cleavable moiety (CM1) coupled tothe AB1, wherein the CM1 is a polypeptide that functions as a substratefor a protease.
 79. A multispecific activatable antibody that in anactivated state binds the epsilon chain of CD3 (CD3ε) and a secondtarget, the multispecific activatable antibody comprising: at least afirst antibody or an antigen binding fragment thereof (AB1) thatspecifically binds CD3ε; at least a first masking moiety (MM1) thatinhibits the binding of the AB1 to CD3ε when the multispecificactivatable antibody is in an uncleaved state; at least a firstcleavable moiety (CM1) coupled to the AB1, wherein the CM1 is apolypeptide that functions as a substrate for a protease; at least asecond antibody or an antigen binding fragment thereof (AB2) thatspecifically binds a second target or epitope; at least a second maskingmoiety (MM2) that inhibits the binding of the AB2 to the second targetor epitope when the multispecific activatable antibody is in anuncleaved state; and at least a second cleavable moiety (CM2) coupled tothe AB2, wherein the CM2 is a polypeptide that functions as a substratefor a protease.
 80. The multispecific activatable antibody of claim 61,claim 78 or claim 79, wherein the MM1 and/or the MM2 has a dissociationconstant for binding to the AB1 and/or AB2 which is selected from thegroup consisting of: greater than the dissociation constant of the AB1to CD3ε and/or than the dissociation constant of the AB2 to the secondtarget or epitope; equivalent to the dissociation constant of the AB1 toCD3ε and/or than the dissociation constant of the AB2 to the secondtarget or epitope; and less than the dissociation constant of the AB1 toCD3ε and/or than the dissociation constant of the AB2 to the secondtarget or epitope.
 81. The multispecific activatable antibody of claim61, claim 78 or claim 79, wherein the MM1 polypeptide sequence isdifferent from that of CD3ε, the MM2 polypeptide sequence is differentfrom that of the second target or epitope, and wherein the MM1polypeptide sequence and/or the MM2 polypeptide sequence is no more than50% identical to any natural binding partner of AB1 and/or AB2.
 82. Themultispecific activatable antibody of claim 61, claim 78 or claim 79,wherein the MM1 and/or MM2 does not interfere or compete with the AB1for binding to CD3ε and/or with the AB2 for binding to the second targetor epitope when the multispecific activatable antibody is in a cleavedstate.
 83. The multispecific activatable antibody of claim 61, claim 78or claim 79, wherein the MM1 comprises a sequence selected from thegroup consisting of the sequences shown in Table 7 or Table
 8. 84. Themultispecific activatable antibody of claim 61, claim 78 or claim 79,wherein the protease that cleaves CM1 and/or CM2 is produced by a tumorthat is in proximity to cells that express CD3ε in a tissue and/orproduced by a tumor that is co-localized with the CD3ε and/or the secondtarget or epitope in a tissue, and wherein the protease cleaves CM1and/or CM2 in the multispecific activatable antibody when themultispecific activatable antibody is exposed to the protease.
 85. Themultispecific activatable antibody of claim 61, claim 78 or claim 79,wherein at least a portion of the multispecific activatable antibody inthe uncleaved state has the structural arrangement from N-terminus toC-terminus as follows: MM1-CM1-AB1, AB1-CM1-MM1, MM2-CM2-AB2, orAB2-CM2-MM2.
 86. The multispecific activatable antibody of claim 61,claim 78 or claim 79, wherein the multispecific activatable antibodycomprises a linking peptide between the MM1 and the CM1 and/or betweenthe MM2 and the CM2.
 87. The multispecific activatable antibody of claim61, claim 78 or claim 79, wherein the multispecific activatable antibodycomprises a linking peptide between the CM1 and the AB1 and/or betweenthe CM2 and the AB2.
 88. The multispecific activatable antibody of claim61, claim 78 or claim 79, wherein the multispecific activatable antibodycomprises a first linking peptide (LP1) and a second linking peptide(LP2), and wherein the multispecific activatable antibody in theuncleaved state has the structural arrangement from N-terminus toC-terminus as follows: MM1-LP1-CM1-LP2-AB1, AB1-LP2-CM1-LP1-MM1,MM2-LP1-CM2-LP2-AB2, or AB2-LP2-CM2-LP1-MM2.
 89. The multispecificactivatable antibody of claim 88, wherein the two linking peptides neednot be identical to each other.
 90. The multispecific activatableantibody of claim 88, wherein each of LP1 and LP2 is a peptide of about1 to 20 amino acids in length.
 91. The multispecific activatableantibody of claim 61, claim 78 or claim 79, wherein the MM1 and/or theMM2 is a polypeptide of no more than 40 amino acids in length.
 92. Themultispecific activatable antibody of claim 61, claim 78 or claim 79,wherein the CM1 and/or CM2 is a polypeptide of up to 15 amino acids inlength.
 93. The multispecific activatable antibody of claim 61, claim 78or claim 79, wherein the CM1 and/or CM2 is a substrate for a proteaseselected from the group consisting of the proteases listed in Table 3 orwherein the CM1 and/or CM2 comprises an amino acid sequence selectedfrom the group consisting of SEQ ID NO: 67-86, 321-341, and 896-926. 94.The multispecific activatable antibody of claim 61, claim 78 or claim79, wherein the antigen binding fragment thereof of AB1 and/or AB2 isselected from the group consisting of a Fab fragment, a F(ab′)₂fragment, a scFv, a scab, a dAb, a single domain heavy chain antibody,and a single domain light chain antibody.
 95. The multispecificactivatable antibody as in any of claims 78 to 94, wherein AB1 comprisesa VH CDR1 comprising the amino acid sequence TYAMN (SEQ ID NO: 53); a VHCD2 comprising the amino acid sequence RIRSKYNNYATYYADSVKD (SEQ ID NO:54); a VH CDR3 comprising the amino acid sequence HGNFGNSYVSWFAY (SEQ IDNO: 55), a VL CDR1 comprising the amino acid sequence RSSTGAVTTSNYAN(SEQ ID NO: 56); a VL CDR2 comprising the amino acid sequence GTNKRAP(SEQ ID NO: 57); and a VL CDR3 comprising the amino acid sequenceALWYSNLWV (SEQ ID NO: 58).
 96. The multispecific activatable antibody asin any of claims 78 to 94, wherein AB1 comprises a variable heavy chain(Hv) comprising the amino acid sequence of SEQ ID NO: 4 and a variablelight chain (Lv) comprising the amino acid sequence of SEQ ID NO:
 2. 97.The multispecific activatable antibody as in any of claims 78 to 94,wherein the AB1 and/or AB2 comprises a scFv fragment.
 98. Themultispecific activatable antibody as in any of claims 78 to 94, whereinthe AB1 comprises a scFv fragment, and wherein the scFv fragmentcomprises the amino acid sequence of SEQ ID NO: 6 or the amino acidsequence of SEQ ID NO:
 30. 99. The multispecific activatable antibody asin any of claims 78 to 94, wherein the AB1 comprises the amino acidsequence of SEQ ID NO:
 587. 100. The multispecific activatable antibodyas in any of claims 78 to 94, wherein the AB1 comprises the amino acidsequence of SEQ ID NO:
 588. 101. The multispecific activatable antibodyas in any of claims 78 to 94, wherein the target of AB2 is selected fromthe group of targets listed in Table
 1. 102. The multispecificactivatable antibody as in any of claims 78 to 94, wherein AB2 is or isderived from an antibody selected from the group of antibodies listed inTable
 2. 103. The multispecific activatable antibody of claim 78 orclaim 79 comprising an agent conjugated to at least one of AB1 and AB2.104. The multispecific activatable antibody of claim 103, wherein theagent is a therapeutic agent, an antineoplastic agent, a toxin orfragment thereof, a detectable moiety or a diagnostic agent.
 105. Themultispecific activatable antibody of claim 103, wherein the agent isconjugated to at least one of AB1 and AB2 via a linker.
 106. Apharmaceutical composition comprising the multispecific antibody as inany of claim 49-55 or 60-77 or the multispecific activatable antibody asin any of claims 78-102 and a carrier.
 107. A pharmaceutical compositioncomprising the multispecific antibody as in any of claims 49-77 or themultispecific activatable antibody as in any of claims 78-102 and acarrier.
 108. An isolated nucleic acid molecule encoding at least aportion of the multispecific antibody as in any of claims 49-54.
 109. Anisolated nucleic acid molecule encoding at least a portion of themultispecific activatable antibody as in any of claims 95-100.
 110. Avector comprising the isolated nucleic acid molecule of claim 108 orclaim
 109. 111. A method of producing a multispecific antibody or amultispecific activatable antibody that specifically binds at least theepsilon chain of CD3 (CD3ε), the method comprising culturing a cellunder conditions that lead to expression of the multispecific antibodyor multispecific activatable antibody, wherein the cell comprises thenucleic acid molecule of claim 108 or claim
 109. 112. A method ofmanufacturing a multispecific activatable antibody that in an activatedstate binds the epsilon chain of CD3 (CD3ε) and a second target, themethod comprising: (a) culturing a cell comprising a nucleic acidconstruct that encodes the multispecific activatable antibody underconditions that lead to expression of the multispecific activatableantibody, wherein the activatable antibody is selected from the groupconsisting of: (i) an activatable antibody comprising at least a firstantibody or an antigen binding fragment thereof (AB1) that specificallybinds CD3ε, a second antibody or an antigen binding fragment thereof(AB2) that specifically binds a second target or epitope, at least afirst masking moiety (MM1) that inhibits the binding of the AB1 to CD3εwhen the multispecific activatable antibody is in an uncleaved state,and at least a first cleavable moiety (CM1) coupled to the AB1, and (ii)an activatable antibody comprising at least a first antibody or anantigen binding fragment thereof (AB1) that specifically binds CD3ε, asecond antibody or an antigen binding fragment thereof (AB2) thatspecifically binds a second target or epitope, a first masking moiety(MM1) that inhibits the binding of the AB1 to CD3ε when themultispecific activatable antibody is in an uncleaved state, a secondmasking moiety (MM2) that inhibits the binding of the AB2 to its targetwhen the multispecific activatable antibody is in an uncleaved state, afirst cleavable moiety (CM1) coupled to the AB1, and a second cleavablemoiety (CM2) coupled to the AB2; and (b) recovering the activatableantibody.
 113. The method of claim 112, wherein the CM1, the CM2, orboth CM1 and CM2 is a polypeptide that functions as a substrate for aprotease.
 114. The method of claim 112, wherein the CM1 is positioned inthe activatable antibody such that, when the activatable antibody is inan uncleaved state, the MM1 interferes with specific binding of the AB1to CD3ε and when the activatable antibody is in a cleaved state the MM1does not interfere or compete with specific binding of the AB1 to CD3ε,and wherein the CM2 is positioned in the activatable antibody such that,when the activatable antibody is in an uncleaved state, the MM2interferes with specific binding of the AB2 to its target and when theactivatable antibody is in a cleaved state the MM2 does not interfere orcompete with specific binding of the AB2 to its target.
 115. A method ofalleviating a symptom of a clinical indication associated with adisorder in a subject, the method comprising administering the isolatedsingle chain variable antibody as in any of claims 1-5 or theactivatable antibody as in any of claim 11-33 or 37-39, or themultispecific antibody as in any of claim 49-54 or 60, or themultispecific activatable antibody as in any of claims 61-102 to asubject in need thereof in an amount sufficient to alleviate the symptomof the clinical indication associated with the disorder.
 116. A methodof alleviating a symptom of a clinical indication associated with adisorder in a subject, the method comprising administering the isolatedsingle chain variable antibody as in any of claims 1-8 or theactivatable antibody as in any of claims 11-39, or the multispecificantibody as in any of claims 49-60, or the multispecific activatableantibody as in any of claims 61-105 to a subject in need thereof in anamount sufficient to alleviate the symptom of the clinical indicationassociated with the disorder.
 117. The method of claim 115 or claim 116,wherein said subject is a human.
 118. The method of claim 115 or claim116, wherein the disorder is cancer.